KR100656888B1 - Composition Comprising Ginsenosides Having Neuroprotective Effects against Brain Striatum Neurotoxicity - Google Patents
Composition Comprising Ginsenosides Having Neuroprotective Effects against Brain Striatum Neurotoxicity Download PDFInfo
- Publication number
- KR100656888B1 KR100656888B1 KR1020040042590A KR20040042590A KR100656888B1 KR 100656888 B1 KR100656888 B1 KR 100656888B1 KR 1020040042590 A KR1020040042590 A KR 1020040042590A KR 20040042590 A KR20040042590 A KR 20040042590A KR 100656888 B1 KR100656888 B1 KR 100656888B1
- Authority
- KR
- South Korea
- Prior art keywords
- red ginseng
- huntington
- neurotoxicity
- delete delete
- ginseng saponin
- Prior art date
Links
- 239000000203 mixture Substances 0.000 title claims abstract description 45
- 229930182494 ginsenoside Natural products 0.000 title claims abstract description 17
- 206010029350 Neurotoxicity Diseases 0.000 title claims abstract description 9
- 206010044221 Toxic encephalopathy Diseases 0.000 title claims abstract description 9
- 230000007135 neurotoxicity Effects 0.000 title claims abstract description 9
- 231100000228 neurotoxicity Toxicity 0.000 title claims abstract description 9
- 210000001577 neostriatum Anatomy 0.000 title abstract description 17
- 210000004556 brain Anatomy 0.000 title description 12
- 230000000324 neuroprotective effect Effects 0.000 title description 7
- WBLZUCOIBUDNBV-UHFFFAOYSA-N 3-nitropropanoic acid Chemical compound OC(=O)CC[N+]([O-])=O WBLZUCOIBUDNBV-UHFFFAOYSA-N 0.000 claims abstract description 68
- 235000002789 Panax ginseng Nutrition 0.000 claims abstract description 47
- 229930182490 saponin Natural products 0.000 claims abstract description 38
- 150000007949 saponins Chemical class 0.000 claims abstract description 38
- 239000001397 quillaja saponaria molina bark Substances 0.000 claims abstract description 37
- 208000023105 Huntington disease Diseases 0.000 claims abstract description 18
- 230000003834 intracellular effect Effects 0.000 claims abstract description 11
- 208000010877 cognitive disease Diseases 0.000 claims abstract description 9
- 208000022821 personality disease Diseases 0.000 claims abstract description 9
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 claims abstract description 8
- 229910001424 calcium ion Inorganic materials 0.000 claims abstract description 8
- 230000006676 mitochondrial damage Effects 0.000 claims abstract description 6
- 230000002265 prevention Effects 0.000 claims abstract description 6
- 210000002161 motor neuron Anatomy 0.000 claims abstract description 5
- 230000004693 neuron damage Effects 0.000 claims abstract description 5
- 208000019430 Motor disease Diseases 0.000 claims abstract description 3
- 230000001149 cognitive effect Effects 0.000 claims abstract description 3
- 230000000694 effects Effects 0.000 claims description 14
- 208000028698 Cognitive impairment Diseases 0.000 claims description 4
- 235000015872 dietary supplement Nutrition 0.000 claims description 4
- 208000028389 Nerve injury Diseases 0.000 claims 1
- 230000006735 deficit Effects 0.000 claims 1
- 230000008764 nerve damage Effects 0.000 claims 1
- 208000024891 symptom Diseases 0.000 abstract description 8
- 208000016285 Movement disease Diseases 0.000 abstract description 6
- 239000003814 drug Substances 0.000 abstract description 6
- 229940089161 ginsenoside Drugs 0.000 abstract description 6
- 239000004480 active ingredient Substances 0.000 abstract 1
- 229940124597 therapeutic agent Drugs 0.000 abstract 1
- 235000017709 saponins Nutrition 0.000 description 34
- 210000004027 cell Anatomy 0.000 description 19
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 13
- 230000003902 lesion Effects 0.000 description 13
- OFOBLEOULBTSOW-UHFFFAOYSA-N Malonic acid Chemical compound OC(=O)CC(O)=O OFOBLEOULBTSOW-UHFFFAOYSA-N 0.000 description 12
- 238000010171 animal model Methods 0.000 description 12
- 241001465754 Metazoa Species 0.000 description 11
- 239000011780 sodium chloride Substances 0.000 description 11
- 241000208340 Araliaceae Species 0.000 description 10
- 235000008434 ginseng Nutrition 0.000 description 10
- 235000005035 Panax pseudoginseng ssp. pseudoginseng Nutrition 0.000 description 9
- 235000003140 Panax quinquefolius Nutrition 0.000 description 9
- 239000011575 calcium Substances 0.000 description 8
- 210000002569 neuron Anatomy 0.000 description 8
- 239000000243 solution Substances 0.000 description 8
- 238000010186 staining Methods 0.000 description 8
- 241000700159 Rattus Species 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- LNEPOXFFQSENCJ-UHFFFAOYSA-N haloperidol Chemical compound C1CC(O)(C=2C=CC(Cl)=CC=2)CCN1CCCC(=O)C1=CC=C(F)C=C1 LNEPOXFFQSENCJ-UHFFFAOYSA-N 0.000 description 6
- 230000002829 reductive effect Effects 0.000 description 5
- 230000004083 survival effect Effects 0.000 description 5
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 4
- 239000007995 HEPES buffer Substances 0.000 description 4
- ZHAFUINZIZIXFC-UHFFFAOYSA-N [9-(dimethylamino)-10-methylbenzo[a]phenoxazin-5-ylidene]azanium;chloride Chemical compound [Cl-].O1C2=CC(=[NH2+])C3=CC=CC=C3C2=NC2=C1C=C(N(C)C)C(C)=C2 ZHAFUINZIZIXFC-UHFFFAOYSA-N 0.000 description 4
- 230000003833 cell viability Effects 0.000 description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 4
- 229940079593 drug Drugs 0.000 description 4
- 238000001647 drug administration Methods 0.000 description 4
- 235000013305 food Nutrition 0.000 description 4
- 210000001700 mitochondrial membrane Anatomy 0.000 description 4
- 210000004498 neuroglial cell Anatomy 0.000 description 4
- 230000001681 protective effect Effects 0.000 description 4
- 210000000130 stem cell Anatomy 0.000 description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 3
- VREFGVBLTWBCJP-UHFFFAOYSA-N alprazolam Chemical compound C12=CC(Cl)=CC=C2N2C(C)=NN=C2CN=C1C1=CC=CC=C1 VREFGVBLTWBCJP-UHFFFAOYSA-N 0.000 description 3
- 210000001130 astrocyte Anatomy 0.000 description 3
- 235000013361 beverage Nutrition 0.000 description 3
- 230000030833 cell death Effects 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 230000006378 damage Effects 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 235000020710 ginseng extract Nutrition 0.000 description 3
- TXEWRVNOAJOINC-UHFFFAOYSA-N ginsenoside Rb2 Natural products CC(=CCCC(OC1OC(COC2OCC(O)C(O)C2O)C(O)C(O)C1O)C3CCC4(C)C3C(O)CC5C6(C)CCC(OC7OC(CO)C(O)C(O)C7OC8OC(CO)C(O)C(O)C8O)C(C)(C)C6CCC45C)C TXEWRVNOAJOINC-UHFFFAOYSA-N 0.000 description 3
- 229960003878 haloperidol Drugs 0.000 description 3
- 230000002055 immunohistochemical effect Effects 0.000 description 3
- 238000001727 in vivo Methods 0.000 description 3
- 238000007912 intraperitoneal administration Methods 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 239000002609 medium Substances 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- BOLDJAUMGUJJKM-LSDHHAIUSA-N renifolin D Natural products CC(=C)[C@@H]1Cc2c(O)c(O)ccc2[C@H]1CC(=O)c3ccc(O)cc3O BOLDJAUMGUJJKM-LSDHHAIUSA-N 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- RWXIFXNRCLMQCD-JBVRGBGGSA-N (20S)-ginsenoside Rg3 Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@H]1CC[C@]2(C)[C@H]3C[C@@H](O)[C@H]4[C@@]([C@@]3(CC[C@H]2C1(C)C)C)(C)CC[C@@H]4[C@@](C)(O)CCC=C(C)C)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O RWXIFXNRCLMQCD-JBVRGBGGSA-N 0.000 description 2
- PKDBCJSWQUOKDO-UHFFFAOYSA-M 2,3,5-triphenyltetrazolium chloride Chemical compound [Cl-].C1=CC=CC=C1C(N=[N+]1C=2C=CC=CC=2)=NN1C1=CC=CC=C1 PKDBCJSWQUOKDO-UHFFFAOYSA-M 0.000 description 2
- FBFMBWCLBGQEBU-GYMUUCMZSA-N 20-gluco-ginsenoside-Rf Natural products O([C@](CC/C=C(\C)/C)(C)[C@@H]1[C@H]2[C@H](O)C[C@H]3[C@](C)([C@]2(C)CC1)C[C@H](O[C@@H]1[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O2)[C@@H](O)[C@H](O)[C@@H](CO)O1)[C@H]1C(C)(C)[C@@H](O)CC[C@]31C)[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 FBFMBWCLBGQEBU-GYMUUCMZSA-N 0.000 description 2
- KJDSORYAHBAGPP-UHFFFAOYSA-N 4-(3,4-diaminophenyl)benzene-1,2-diamine;hydron;tetrachloride Chemical compound Cl.Cl.Cl.Cl.C1=C(N)C(N)=CC=C1C1=CC=C(N)C(N)=C1 KJDSORYAHBAGPP-UHFFFAOYSA-N 0.000 description 2
- 206010002091 Anaesthesia Diseases 0.000 description 2
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 2
- AGBCLJAHARWNLA-DQUQINEDSA-N Ginsenoside Rg2 Natural products O[C@@H]1[C@H](O)[C@@H](O)[C@H](C)O[C@H]1O[C@H]1[C@H](O[C@@H]2[C@H]3C(C)(C)[C@@H](O)CC[C@]3(C)[C@@H]3[C@@]([C@@]4(CC[C@@H]([C@H]4[C@H](O)C3)[C@@](C)(O)CCC=C(C)C)C)(C)C2)O[C@H](CO)[C@@H](O)[C@@H]1O AGBCLJAHARWNLA-DQUQINEDSA-N 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 description 2
- AFCARXCZXQIEQB-UHFFFAOYSA-N N-[3-oxo-3-(2,4,6,7-tetrahydrotriazolo[4,5-c]pyridin-5-yl)propyl]-2-[[3-(trifluoromethoxy)phenyl]methylamino]pyrimidine-5-carboxamide Chemical compound O=C(CCNC(=O)C=1C=NC(=NC=1)NCC1=CC(=CC=C1)OC(F)(F)F)N1CC2=C(CC1)NN=N2 AFCARXCZXQIEQB-UHFFFAOYSA-N 0.000 description 2
- ACFIXJIJDZMPPO-NNYOXOHSSA-N NADPH Chemical compound C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](OP(O)(O)=O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 ACFIXJIJDZMPPO-NNYOXOHSSA-N 0.000 description 2
- 229930040373 Paraformaldehyde Natural products 0.000 description 2
- 206010034972 Photosensitivity reaction Diseases 0.000 description 2
- YURJSTAIMNSZAE-UHFFFAOYSA-N UNPD89172 Natural products C1CC(C2(CC(C3C(C)(C)C(O)CCC3(C)C2CC2O)OC3C(C(O)C(O)C(CO)O3)O)C)(C)C2C1C(C)(CCC=C(C)C)OC1OC(CO)C(O)C(O)C1O YURJSTAIMNSZAE-UHFFFAOYSA-N 0.000 description 2
- 230000001154 acute effect Effects 0.000 description 2
- 230000037005 anaesthesia Effects 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 230000003542 behavioural effect Effects 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 230000001684 chronic effect Effects 0.000 description 2
- 239000003210 dopamine receptor blocking agent Substances 0.000 description 2
- 239000002552 dosage form Substances 0.000 description 2
- 230000005284 excitation Effects 0.000 description 2
- YFHXZQPUBCBNIP-UHFFFAOYSA-N fura-2 Chemical compound CC1=CC=C(N(CC(O)=O)CC(O)=O)C(OCCOC=2C(=CC=3OC(=CC=3C=2)C=2OC(=CN=2)C(O)=O)N(CC(O)=O)CC(O)=O)=C1 YFHXZQPUBCBNIP-UHFFFAOYSA-N 0.000 description 2
- YURJSTAIMNSZAE-HHNZYBFYSA-N ginsenoside Rg1 Chemical compound O([C@@](C)(CCC=C(C)C)[C@@H]1[C@@H]2[C@@]([C@@]3(C[C@@H]([C@H]4C(C)(C)[C@@H](O)CC[C@]4(C)[C@H]3C[C@H]2O)O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)O)C)(C)CC1)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O YURJSTAIMNSZAE-HHNZYBFYSA-N 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- 210000001320 hippocampus Anatomy 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- 230000005764 inhibitory process Effects 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 230000004065 mitochondrial dysfunction Effects 0.000 description 2
- 239000003068 molecular probe Substances 0.000 description 2
- VMGAPWLDMVPYIA-HIDZBRGKSA-N n'-amino-n-iminomethanimidamide Chemical compound N\N=C\N=N VMGAPWLDMVPYIA-HIDZBRGKSA-N 0.000 description 2
- 230000001537 neural effect Effects 0.000 description 2
- 229930027945 nicotinamide-adenine dinucleotide Natural products 0.000 description 2
- 229920002866 paraformaldehyde Polymers 0.000 description 2
- 239000008363 phosphate buffer Substances 0.000 description 2
- 208000007578 phototoxic dermatitis Diseases 0.000 description 2
- 231100000018 phototoxicity Toxicity 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 238000010998 test method Methods 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- 229910052724 xenon Inorganic materials 0.000 description 2
- FHNFHKCVQCLJFQ-UHFFFAOYSA-N xenon atom Chemical compound [Xe] FHNFHKCVQCLJFQ-UHFFFAOYSA-N 0.000 description 2
- AGBCLJAHARWNLA-UHFFFAOYSA-N (20R)-ginsenoside Rg2 Natural products OC1C(O)C(O)C(C)OC1OC1C(OC2C3C(C)(C)C(O)CCC3(C)C3C(C4(CCC(C4C(O)C3)C(C)(O)CCC=C(C)C)C)(C)C2)OC(CO)C(O)C1O AGBCLJAHARWNLA-UHFFFAOYSA-N 0.000 description 1
- FBFMBWCLBGQEBU-RXMALORBSA-N (2s,3r,4s,5s,6r)-2-[(2r,3r,4s,5s,6r)-2-[[(3s,5r,6s,8r,9r,10r,12r,13r,14r,17s)-3,12-dihydroxy-4,4,8,10,14-pentamethyl-17-[(2s)-6-methyl-2-[(2s,3r,4s,5s,6r)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxyhept-5-en-2-yl]-2,3,5,6,7,9,11,12,13,15,16,17-dodecah Chemical compound O([C@@](C)(CCC=C(C)C)[C@@H]1[C@@H]2[C@@]([C@@]3(C[C@@H]([C@H]4C(C)(C)[C@@H](O)CC[C@]4(C)[C@H]3C[C@H]2O)O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)O)C)(C)CC1)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O FBFMBWCLBGQEBU-RXMALORBSA-N 0.000 description 1
- DNXIKVLOVZVMQF-UHFFFAOYSA-N (3beta,16beta,17alpha,18beta,20alpha)-17-hydroxy-11-methoxy-18-[(3,4,5-trimethoxybenzoyl)oxy]-yohimban-16-carboxylic acid, methyl ester Natural products C1C2CN3CCC(C4=CC=C(OC)C=C4N4)=C4C3CC2C(C(=O)OC)C(O)C1OC(=O)C1=CC(OC)=C(OC)C(OC)=C1 DNXIKVLOVZVMQF-UHFFFAOYSA-N 0.000 description 1
- NFZYDZXHKFHPGA-UHFFFAOYSA-N 17alpha-hydroxygofruside Natural products C12CC(C)(C)CCC2(C(=O)OC2C(C(O)C(O)C(CO)O2)O)CCC(C2(CCC3C4(C)C)C)(C)C1=CCC2C3(C)CCC4OC1OC(C(O)=O)C(O)C(O)C1OC1OC(CO)C(O)C(O)C1O NFZYDZXHKFHPGA-UHFFFAOYSA-N 0.000 description 1
- QVGLDPPIMKSVBG-UHFFFAOYSA-N 2-methylbutane Chemical compound CCC(C)C.CCC(C)C QVGLDPPIMKSVBG-UHFFFAOYSA-N 0.000 description 1
- AZKSAVLVSZKNRD-UHFFFAOYSA-M 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide Chemical compound [Br-].S1C(C)=C(C)N=C1[N+]1=NC(C=2C=CC=CC=2)=NN1C1=CC=CC=C1 AZKSAVLVSZKNRD-UHFFFAOYSA-M 0.000 description 1
- 229930000680 A04AD01 - Scopolamine Natural products 0.000 description 1
- 206010000117 Abnormal behaviour Diseases 0.000 description 1
- 101800000263 Acidic protein Proteins 0.000 description 1
- 206010001497 Agitation Diseases 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 208000000044 Amnesia Diseases 0.000 description 1
- 206010003694 Atrophy Diseases 0.000 description 1
- 239000012583 B-27 Supplement Substances 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 208000014644 Brain disease Diseases 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- 102000011727 Caspases Human genes 0.000 description 1
- 108010076667 Caspases Proteins 0.000 description 1
- 206010008096 Cerebral atrophy Diseases 0.000 description 1
- QXQFFGOMXYKNBA-UHFFFAOYSA-N Chikusetsusaponin V Natural products CC1(C)CCC2(CCC3C(=CCC4C3(C)CCC5C(C)(C)C(CCC45C)OC6OC(C(O)C(O)C6OC7OC(CO)C(O)C(O)C7O)C(=O)O)C2C1)C(=O)OC8OC(CO)C(O)C(O)C8O QXQFFGOMXYKNBA-UHFFFAOYSA-N 0.000 description 1
- NFZYDZXHKFHPGA-QQHDHSITSA-N Chikusetsusaponin-V Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@H](O[C@H]1O[C@H]1CC[C@]2(C)[C@H]3CC=C4[C@@]([C@@]3(CC[C@H]2C1(C)C)C)(C)CC[C@]1(CCC(C[C@H]14)(C)C)C(=O)O[C@H]1[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O1)O)C(O)=O)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O NFZYDZXHKFHPGA-QQHDHSITSA-N 0.000 description 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
- 230000007018 DNA scission Effects 0.000 description 1
- 101710088194 Dehydrogenase Proteins 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- 241001269524 Dura Species 0.000 description 1
- 208000012661 Dyskinesia Diseases 0.000 description 1
- 208000000059 Dyspnea Diseases 0.000 description 1
- 206010013975 Dyspnoeas Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- UFNDONGOJKNAES-UHFFFAOYSA-N Ginsenoside Rb1 Natural products CC(=CCCC(C)(OC1OC(COC2OC(CO)C(O)C(O)C2O)C(O)C(O)C1O)C3CCC4(C)C3C(O)CC5C6(C)CCC(OC7OC(CO)C(O)C(O)C7OC8OC(CO)C(O)C(O)C8O)C(C)(C)C6CC(O)C45C)C UFNDONGOJKNAES-UHFFFAOYSA-N 0.000 description 1
- HYPFYJBWSTXDAS-UHFFFAOYSA-N Ginsenoside Rd Natural products CC(=CCCC(C)(OC1OC(CO)C(O)C(O)C1O)C2CCC3(C)C4CCC5C(C)(C)C(CCC5(C)C4CC(O)C23C)OC6OC(CO)C(O)C(O)C6OC7OC(CO)C(O)C(O)C7O)C HYPFYJBWSTXDAS-UHFFFAOYSA-N 0.000 description 1
- UZIOUZHBUYLDHW-MSJHMJQNSA-N Ginsenoside Rf Natural products O([C@H]1[C@@H](O)[C@H](O)[C@H](CO)O[C@@H]1O[C@@H]1[C@H]2C(C)(C)[C@@H](O)CC[C@]2(C)[C@@H]2[C@](C)([C@@]3(C)[C@H]([C@@H](O)C2)[C@@H]([C@@](O)(CC/C=C(\C)/C)C)CC3)C1)[C@H]1[C@@H](O)[C@@H](O)[C@H](O)[C@H](CO)O1 UZIOUZHBUYLDHW-MSJHMJQNSA-N 0.000 description 1
- XIRZPICFRDZXPF-UHFFFAOYSA-N Ginsenoside Rg3 Natural products CC(C)=CCCC(C)(O)C1CCC(C2(CC(O)C3C4(C)C)C)(C)C1C(O)CC2C3(C)CCC4OC1OC(CO)C(O)C(O)C1OC1OC(CO)C(O)C(O)C1O XIRZPICFRDZXPF-UHFFFAOYSA-N 0.000 description 1
- 239000012981 Hank's balanced salt solution Substances 0.000 description 1
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 description 1
- STECJAGHUSJQJN-GAUPFVANSA-N Hyoscine Natural products C1([C@H](CO)C(=O)OC2C[C@@H]3N([C@H](C2)[C@@H]2[C@H]3O2)C)=CC=CC=C1 STECJAGHUSJQJN-GAUPFVANSA-N 0.000 description 1
- 102000004310 Ion Channels Human genes 0.000 description 1
- 208000026139 Memory disease Diseases 0.000 description 1
- 101000888418 Mus musculus Glial fibrillary acidic protein Proteins 0.000 description 1
- STECJAGHUSJQJN-UHFFFAOYSA-N N-Methyl-scopolamin Natural products C1C(C2C3O2)N(C)C3CC1OC(=O)C(CO)C1=CC=CC=C1 STECJAGHUSJQJN-UHFFFAOYSA-N 0.000 description 1
- 208000012902 Nervous system disease Diseases 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 240000004371 Panax ginseng Species 0.000 description 1
- 206010033799 Paralysis Diseases 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 206010034719 Personality change Diseases 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- LCQMZZCPPSWADO-UHFFFAOYSA-N Reserpilin Natural products COC(=O)C1COCC2CN3CCc4c([nH]c5cc(OC)c(OC)cc45)C3CC12 LCQMZZCPPSWADO-UHFFFAOYSA-N 0.000 description 1
- QEVHRUUCFGRFIF-SFWBKIHZSA-N Reserpine Natural products O=C(OC)[C@@H]1[C@H](OC)[C@H](OC(=O)c2cc(OC)c(OC)c(OC)c2)C[C@H]2[C@@H]1C[C@H]1N(C2)CCc2c3c([nH]c12)cc(OC)cc3 QEVHRUUCFGRFIF-SFWBKIHZSA-N 0.000 description 1
- 240000000111 Saccharum officinarum Species 0.000 description 1
- 235000007201 Saccharum officinarum Nutrition 0.000 description 1
- UIIMBOGNXHQVGW-DEQYMQKBSA-M Sodium bicarbonate-14C Chemical compound [Na+].O[14C]([O-])=O UIIMBOGNXHQVGW-DEQYMQKBSA-M 0.000 description 1
- 206010041349 Somnolence Diseases 0.000 description 1
- 238000000692 Student's t-test Methods 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229920004890 Triton X-100 Polymers 0.000 description 1
- 239000013504 Triton X-100 Substances 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 230000005856 abnormality Effects 0.000 description 1
- 230000001133 acceleration Effects 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 238000000540 analysis of variance Methods 0.000 description 1
- 230000037444 atrophy Effects 0.000 description 1
- 210000004227 basal ganglia Anatomy 0.000 description 1
- 230000006399 behavior Effects 0.000 description 1
- 230000006741 behavioral dysfunction Effects 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- 239000001569 carbon dioxide Substances 0.000 description 1
- 229910002092 carbon dioxide Inorganic materials 0.000 description 1
- 210000001159 caudate nucleus Anatomy 0.000 description 1
- 230000005779 cell damage Effects 0.000 description 1
- 208000037887 cell injury Diseases 0.000 description 1
- 230000006727 cell loss Effects 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 210000003169 central nervous system Anatomy 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- CNHRRMQBWQJRPN-UHFFFAOYSA-N chikusetsusaponin LM5 Natural products C1CC(C2(CC(O)C3C(C)(C)C(OC4C(C(O)C(O)C(CO)O4)OC4C(C(O)C(O)C(CO)O4)O)CCC3(C)C2CC2O)C)(C)C2C1C(C)(CCC=C(C)C)OC(C(C(O)C1O)O)OC1COC1OC(CO)C(O)C1O CNHRRMQBWQJRPN-UHFFFAOYSA-N 0.000 description 1
- 239000000460 chlorine Substances 0.000 description 1
- 229910052801 chlorine Inorganic materials 0.000 description 1
- QZUDBNBUXVUHMW-UHFFFAOYSA-N clozapine Chemical compound C1CN(C)CCN1C1=NC2=CC(Cl)=CC=C2NC2=CC=CC=C12 QZUDBNBUXVUHMW-UHFFFAOYSA-N 0.000 description 1
- 229960004170 clozapine Drugs 0.000 description 1
- 230000015271 coagulation Effects 0.000 description 1
- 238000005345 coagulation Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 210000003618 cortical neuron Anatomy 0.000 description 1
- 231100000135 cytotoxicity Toxicity 0.000 description 1
- 230000003013 cytotoxicity Effects 0.000 description 1
- 238000007405 data analysis Methods 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 230000007850 degeneration Effects 0.000 description 1
- 230000003412 degenerative effect Effects 0.000 description 1
- 235000005911 diet Nutrition 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 231100000676 disease causative agent Toxicity 0.000 description 1
- 208000035475 disorder Diseases 0.000 description 1
- 230000008451 emotion Effects 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- ZMMJGEGLRURXTF-UHFFFAOYSA-N ethidium bromide Chemical compound [Br-].C12=CC(N)=CC=C2C2=CC=C(N)C=C2[N+](CC)=C1C1=CC=CC=C1 ZMMJGEGLRURXTF-UHFFFAOYSA-N 0.000 description 1
- 229960005542 ethidium bromide Drugs 0.000 description 1
- 239000000834 fixative Substances 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 238000013467 fragmentation Methods 0.000 description 1
- 238000006062 fragmentation reaction Methods 0.000 description 1
- 210000001652 frontal lobe Anatomy 0.000 description 1
- 235000013376 functional food Nutrition 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- GZYPWOGIYAIIPV-JBDTYSNRSA-N ginsenoside Rb1 Chemical compound C([C@H]1O[C@H]([C@@H]([C@@H](O)[C@@H]1O)O)O[C@@](C)(CCC=C(C)C)[C@@H]1[C@@H]2[C@@]([C@@]3(CC[C@H]4C(C)(C)[C@@H](O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O)CC[C@]4(C)[C@H]3C[C@H]2O)C)(C)CC1)O[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O GZYPWOGIYAIIPV-JBDTYSNRSA-N 0.000 description 1
- NODILNFGTFIURN-GZPRDHCNSA-N ginsenoside Rb2 Chemical compound C([C@H]1O[C@H]([C@@H]([C@@H](O)[C@@H]1O)O)O[C@@](C)(CCC=C(C)C)[C@@H]1[C@@H]2[C@@]([C@@]3(CC[C@H]4C(C)(C)[C@@H](O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O)CC[C@]4(C)[C@H]3C[C@H]2O)C)(C)CC1)O[C@@H]1OC[C@H](O)[C@H](O)[C@H]1O NODILNFGTFIURN-GZPRDHCNSA-N 0.000 description 1
- PFSIGTQOILYIIU-UHFFFAOYSA-N ginsenoside Rb3 Natural products CC(=CCCC(C)(O)C1CCC2(C)C3CCC4C(C)(C)C(CCC4(C)C3CC(OC5OC(COC6OCC(O)C(O)C6O)C(O)C(O)C5O)C12C)OC7OC(CO)C(O)C(O)C7OC8OC(CO)C(O)C(O)C8O)C PFSIGTQOILYIIU-UHFFFAOYSA-N 0.000 description 1
- JDCPEKQWFDWQLI-LUQKBWBOSA-N ginsenoside Rc Chemical compound C([C@H]1O[C@H]([C@@H]([C@@H](O)[C@@H]1O)O)O[C@@](C)(CCC=C(C)C)[C@@H]1[C@@H]2[C@@]([C@@]3(CC[C@H]4C(C)(C)[C@@H](O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O)CC[C@]4(C)[C@H]3C[C@H]2O)C)(C)CC1)O[C@@H]1O[C@@H](CO)[C@H](O)[C@H]1O JDCPEKQWFDWQLI-LUQKBWBOSA-N 0.000 description 1
- PWAOOJDMFUQOKB-WCZZMFLVSA-N ginsenoside Re Chemical compound O[C@@H]1[C@H](O)[C@@H](O)[C@H](C)O[C@H]1O[C@H]1[C@H](O[C@@H]2[C@H]3C(C)(C)[C@@H](O)CC[C@]3(C)[C@@H]3[C@@]([C@@]4(CC[C@@H]([C@H]4[C@H](O)C3)[C@](C)(CCC=C(C)C)O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O3)O)C)(C)C2)O[C@H](CO)[C@@H](O)[C@@H]1O PWAOOJDMFUQOKB-WCZZMFLVSA-N 0.000 description 1
- UZIOUZHBUYLDHW-XUBRWZAZSA-N ginsenoside Rf Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@H]2C(C)(C)[C@@H](O)CC[C@]2(C)[C@H]2C[C@@H](O)[C@H]3[C@@]([C@@]2(C1)C)(C)CC[C@@H]3[C@@](C)(O)CCC=C(C)C)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O UZIOUZHBUYLDHW-XUBRWZAZSA-N 0.000 description 1
- CBEHEBUBNAGGKC-UHFFFAOYSA-N ginsenoside Rg1 Natural products CC(=CCCC(C)(OC1OC(CO)C(O)C(O)C1O)C2CCC3(C)C2C(O)CC4C5(C)CCC(O)C(C)(C)C5CC(OC6OC(CO)C(O)C(O)C6O)C34C)C CBEHEBUBNAGGKC-UHFFFAOYSA-N 0.000 description 1
- RBRANZURTULKJD-UHFFFAOYSA-N ginsenoside Ro Natural products CC1(C)CCC2(CCC3(C)C(=CCC4C5(C)CCC(OC6OC(C)(C(O)C(O)C6OC7OC(CO)C(O)C(O)C7O)C(=O)O)C(C)(C)C5CCC34C)C2C1)C(=O)OC8OC(CO)C(O)C(O)C8O RBRANZURTULKJD-UHFFFAOYSA-N 0.000 description 1
- SPFXZQZPHXUJSR-UHFFFAOYSA-N ginsenoside-Rc Natural products CC(=CCCC(C)(OC1OC(CO)C(O)C(O)C1OC2OC(CO)C(O)C2O)C3CCC4(C)C3C(O)CC5C6(C)CCC(OC7OC(CO)C(O)C(O)C7OC8OC(CO)C(O)C(O)C8O)C(C)(C)C6CCC45C)C SPFXZQZPHXUJSR-UHFFFAOYSA-N 0.000 description 1
- ZTQSADJAYQOCDD-UHFFFAOYSA-N ginsenoside-Rd2 Natural products C1CC(C2(CCC3C(C)(C)C(OC4C(C(O)C(O)C(CO)O4)O)CCC3(C)C2CC2O)C)(C)C2C1C(C)(CCC=C(C)C)OC(C(C(O)C1O)O)OC1COC1OCC(O)C(O)C1O ZTQSADJAYQOCDD-UHFFFAOYSA-N 0.000 description 1
- AOGZLQUEBLOQCI-UHFFFAOYSA-N ginsenoside-Re Natural products CC1OC(OCC2OC(OC3CC4(C)C(CC(O)C5C(CCC45C)C(C)(CCC=C(C)C)OC6OC(CO)C(O)C(O)C6O)C7(C)CCC(O)C(C)(C)C37)C(O)C(O)C2O)C(O)C(O)C1O AOGZLQUEBLOQCI-UHFFFAOYSA-N 0.000 description 1
- 230000002518 glial effect Effects 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 229960002897 heparin Drugs 0.000 description 1
- 229920000669 heparin Polymers 0.000 description 1
- 241000411851 herbal medicine Species 0.000 description 1
- 230000001744 histochemical effect Effects 0.000 description 1
- 238000010191 image analysis Methods 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 239000000411 inducer Substances 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 230000002147 killing effect Effects 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000006984 memory degeneration Effects 0.000 description 1
- 239000003695 memory enhancer Substances 0.000 description 1
- 208000023060 memory loss Diseases 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- VHGXRGXCDVQIKS-KRWDZBQOSA-N methyl (2s)-3-(4-methylphenyl)sulfonyloxy-2-(phenylmethoxycarbonylamino)propanoate Chemical compound C([C@@H](C(=O)OC)NC(=O)OCC=1C=CC=CC=1)OS(=O)(=O)C1=CC=C(C)C=C1 VHGXRGXCDVQIKS-KRWDZBQOSA-N 0.000 description 1
- 230000002438 mitochondrial effect Effects 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 208000015122 neurodegenerative disease Diseases 0.000 description 1
- 230000016273 neuron death Effects 0.000 description 1
- 231100000189 neurotoxic Toxicity 0.000 description 1
- 230000002887 neurotoxic effect Effects 0.000 description 1
- 239000002858 neurotransmitter agent Substances 0.000 description 1
- JPXMTWWFLBLUCD-UHFFFAOYSA-N nitro blue tetrazolium(2+) Chemical compound COC1=CC(C=2C=C(OC)C(=CC=2)[N+]=2N(N=C(N=2)C=2C=CC=CC=2)C=2C=CC(=CC=2)[N+]([O-])=O)=CC=C1[N+]1=NC(C=2C=CC=CC=2)=NN1C1=CC=C([N+]([O-])=O)C=C1 JPXMTWWFLBLUCD-UHFFFAOYSA-N 0.000 description 1
- JURZHOVRCOWZFN-UHFFFAOYSA-N notoginsenoside R1 Natural products CC(=CCCC(C)(OC1OC(CO)C(O)C(O)C1O)C2CCC3(C)C2C(O)CC4C5(C)CCC(O)C(C)(C)C5C(CC34C)OC6OC(COC7OCC(O)C(O)C7O)C(O)C(O)C6O)C JURZHOVRCOWZFN-UHFFFAOYSA-N 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 230000036542 oxidative stress Effects 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 230000008447 perception Effects 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 229920001992 poloxamer 407 Polymers 0.000 description 1
- 229920000729 poly(L-lysine) polymer Polymers 0.000 description 1
- -1 polyethylene Polymers 0.000 description 1
- 229920000573 polyethylene Polymers 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000000069 prophylactic effect Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 208000020016 psychiatric disease Diseases 0.000 description 1
- 210000002637 putamen Anatomy 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- BJOIZNZVOZKDIG-MDEJGZGSSA-N reserpine Chemical compound O([C@H]1[C@@H]([C@H]([C@H]2C[C@@H]3C4=C([C]5C=CC(OC)=CC5=N4)CCN3C[C@H]2C1)C(=O)OC)OC)C(=O)C1=CC(OC)=C(OC)C(OC)=C1 BJOIZNZVOZKDIG-MDEJGZGSSA-N 0.000 description 1
- 229960003147 reserpine Drugs 0.000 description 1
- MDMGHDFNKNZPAU-UHFFFAOYSA-N roserpine Natural products C1C2CN3CCC(C4=CC=C(OC)C=C4N4)=C4C3CC2C(OC(C)=O)C(OC)C1OC(=O)C1=CC(OC)=C(OC)C(OC)=C1 MDMGHDFNKNZPAU-UHFFFAOYSA-N 0.000 description 1
- 238000010825 rotarod performance test Methods 0.000 description 1
- STECJAGHUSJQJN-FWXGHANASA-N scopolamine Chemical compound C1([C@@H](CO)C(=O)O[C@H]2C[C@@H]3N([C@H](C2)[C@@H]2[C@H]3O2)C)=CC=CC=C1 STECJAGHUSJQJN-FWXGHANASA-N 0.000 description 1
- 229960002646 scopolamine Drugs 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 239000012679 serum free medium Substances 0.000 description 1
- 239000004017 serum-free culture medium Substances 0.000 description 1
- 238000013223 sprague-dawley female rat Methods 0.000 description 1
- 238000013222 sprague-dawley male rat Methods 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 239000003826 tablet Substances 0.000 description 1
- 210000003478 temporal lobe Anatomy 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 238000001665 trituration Methods 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- UOJAEODBOCLNBU-UHFFFAOYSA-N vinaginsenoside R4 Natural products C1CC(C2(CC(O)C3C(C)(C)C(OC4C(C(O)C(O)C(CO)O4)OC4C(C(O)C(O)C(CO)O4)O)CCC3(C)C2CC2O)C)(C)C2C1C(C)(CCC=C(C)C)OC1OC(CO)C(O)C(O)C1O UOJAEODBOCLNBU-UHFFFAOYSA-N 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 230000003313 weakening effect Effects 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/25—Araliaceae (Ginseng family), e.g. ivy, aralia, schefflera or tetrapanax
- A61K36/258—Panax (ginseng)
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/322—Foods, ingredients or supplements having a functional effect on health having an effect on the health of the nervous system or on mental function
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Biomedical Technology (AREA)
- Neurology (AREA)
- Botany (AREA)
- Neurosurgery (AREA)
- Medicinal Chemistry (AREA)
- Veterinary Medicine (AREA)
- Mycology (AREA)
- Pharmacology & Pharmacy (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Medical Informatics (AREA)
- Epidemiology (AREA)
- Microbiology (AREA)
- Hospice & Palliative Care (AREA)
- Biotechnology (AREA)
- Alternative & Traditional Medicine (AREA)
- Psychiatry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Organic Chemistry (AREA)
- Nutrition Science (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Steroid Compounds (AREA)
Abstract
본 발명은 홍삼 사포닌 혼합물을 유효성분으로 함유하는 인지장애, 운동장애, 성격장애 또는 헌팅톤병의 예방 및 치료용 조성물에 관한 것이다. 본 발명의 홍삼 사포닌 혼합물은 헌팅톤병(Huntington's disease)과 유사한 임상적 증상을 유발시키는 것으로 알려진 3-NP에 의해 유도되는 선조체 신경독성, 운동신경 손상, 세포내 칼슘이온 농도 증가 및 미토콘드리아 손상을 유의적으로 억제하므로 인지장애, 운동장애, 성격장애 및 헌팅톤병에 유용한 치료제로 사용될 수 있다. The present invention relates to a composition for the prevention and treatment of cognitive disorders, movement disorders, personality disorders or Huntington's disease containing red ginseng saponin mixture as an active ingredient. The red ginseng saponin mixture of the present invention significantly reduces striatal neurotoxicity, motor neuron damage, increased intracellular calcium ion concentration and mitochondrial damage induced by 3-NP, which is known to cause clinical symptoms similar to Huntington's disease. It can be used as a therapeutic agent for cognitive, motor, personality and Huntington's disease.
홍삼 추출물, 진세노사이드, 3-니트로프로피온산, 선조체, 신경독성Red ginseng extract, ginsenoside, 3-nitropropionic acid, striatum, neurotoxicity
Description
도 1은 본 발명의 대표적인 8개의 진세노사이드의 화학구조를 나타낸 것이다.1 shows the chemical structures of eight representative ginsenosides of the present invention.
도 2는 3-NP의 복강내 투여에 의해 유도된 선조체 병변에 대한 홍삼 추출물(GE) 및 GTS[(홍삼(Red Ginseng) 전체 사포닌 혼합물]의 신경세포 보호효과를 나타낸 것이다.Figure 2 shows the neuronal protective effect of red ginseng extract (GE) and GTS (Red Ginseng total saponin mixture) against striatal lesions induced by intraperitoneal administration of 3-NP.
도 3은 또 다른 선조체 병변 유발 물질인 말론산의 선조체내 투여에 의해 유도된 선조체 병변에 대한 GTS의 신경세포 보호효과를 나타낸 것이다.Figure 3 shows the neuroprotective effect of GTS on striatum lesions induced by striatum administration of malonic acid, another striatum lesion inducer.
도 4는 식염수 투여군(대조군), 3-NP 투여군 및 GTS +3-NP 투여군에서 실험동물의 관상절편을 크레실 바이올렛(CV), GFAP 및 NADPH-다이포라아제로 염색한 결과를 나타낸 것이다.Figure 4 shows the results of staining the coronal sections of the experimental animals in the saline administration (control), 3-NP administration group and GTS + 3-NP administration group with cresyl violet (CV), GFAP and NADPH-diphorase.
도 5는 3-NP에 의해 유도된 운동능력 상실에 대한 GTS의 운동력 증진 효과를 나타낸 것이다( * P<0.05, GTS +3-NP 투여군과 비교; ** P <0.001, 대조군 또는 GTS +3-NP 투여군과 비교).Figure 5 shows the exercise enhancing effect of GTS on the loss of exercise induced by 3-NP ( * P <0.05, compared with GTS + 3-NP administration group; ** P <0.001, control or GTS + 3- Compared with the NP administration group).
도 6은 3-NP에 의해 유도된 실험동물의 사망률에 대한 GTS의 사망률 억제 및 수명 연장 효과를 나타낸 것이다.Figure 6 shows the effect of GTS mortality inhibition and lifespan on the mortality of 3-NP induced animals.
도 7은 3-NP에 의해 유도된 세포내 칼슘이온 농도증가에 대한 GTS의 억제 효과를 나타낸 것이다.Figure 7 shows the inhibitory effect of GTS on the increase in intracellular calcium ion concentration induced by 3-NP.
도 8은 3-NP에 의해 유도된 뇌 선조체 세포들의 미토콘드리아 세포막의 포텐셜 (mitocondrial membrane potential)변화에 대한 GTS의 효과를 나타낸 것이다.Figure 8 shows the effect of GTS on the change in the potential (mitocondrial membrane potential) of mitochondrial cell membrane of brain progenitor cells induced by 3-NP.
도 9는 3-NP에 의해 유도된 배양된 뇌 선조체 세포들의 사멸에 대한 GTS의 방어 효과를 나타낸 것이다. Figure 9 shows the protective effect of GTS on the killing of cultured brain progenitor cells induced by 3-NP.
도 10은 3-NP에 의해 유도된 배양된 뇌 선조체 세포들의 손상에 의하여 유발된 DNA 절단 현상에 대한 GTS의 방어 효과를 나타낸 것이다. FIG. 10 shows the protective effect of GTS against DNA cleavage caused by damage of cultured brain progenitor cells induced by 3-NP.
본 발명은 홍삼 추출물 또는 진세노사이드를 함유하는 3-니트로프로피온 산(3-nitropropionic acid; 3-NP)에 의한 신경세포의 손상 및 사멸로 인한 질환의 예방 및 치료를 위한 조성물에 관한 것이다.The present invention relates to a composition for the prevention and treatment of diseases caused by damage and death of nerve cells by 3-nitropropionic acid (3-NP) containing red ginseng extract or ginsenoside.
파낙스 진셍(Panax ginseng C.A. Meyer (Araliaceae))의 뿌리인 인삼은 아시아뿐만 아니라 세계적으로 가장 많이 사용되는 생약 중의 하나이다. 이러한 인삼의 생물학적 효능을 갖는 성분은 진세노사이드(ginsenoside) 또는 인삼 사포닌(ginseng saponins)으로 알려진 화합물로 30종 이상의 진세노사이드가 인삼에서 분리되어 그 구조가 알려져 있다(Liu, C.X. et al., J. Ethnopharmacol. 36:27-38, 1996; Baek, N.I. et al., Planta Med., 62:86-87, 1996). 그 대표적 예로 진세노사이드-Ra, -Rb1, -Rb2, -Rc, -Rd, -Re, Rf, -Rg1, -Rg2, -Rg3, -Rh1, -Rh2 등이 있다. Ginseng, the root of Panax ginseng CA Meyer ( Araliaceae ), is one of the most widely used herbals in Asia as well as worldwide. The biologically effective component of ginseng is a compound known as ginsenoside or ginseng saponins. More than 30 kinds of ginsenosides are separated from ginseng and its structure is known (Liu, CX et al. , J. Ethnopharmacol. 36: 27-38, 1996; Baek, NI et al. , Planta Med., 62: 86-87, 1996). Representative examples thereof include ginsenoside-Ra, -Rb1, -Rb2, -Rc, -Rd, -Re, Rf, -Rg1, -Rg2, -Rg3, -Rh1, -Rh2 and the like.
최근에는 생체내의 진세노사이드의 다양한 효능 중 중추신경에 대한 효능들이 보고되고 있다. 학습과 기억력에 대한 동물모델에서 인삼의 학습 및 기억력에 대한 효과가 밝혀지고 있으며, 특히 스코폴아민(scopolamine)으로 손상된 동물모델이나 나이에 따라 약해지는 기억력, 기억형성에 가장 중요한 부위로 알려진 해마(hippocampus) 신경세포의 손상에 의한 기억력 감퇴를 인삼이 완화시키는 것으로 알려지고 있다(Hsieh, M.T. et al., Phytother. Res., 14:375-377, 2000; Wesnes, K.A., Ward et al., Psychopharmacology(Berl), 152:353-361, 2000). 인삼의 이러한 기억력에 대한 효과는 인삼 추출물을 포함하는 기억력 증진제로 출원된 바 있다(한국 특허 출원 제2000-4715호). 또한, 다양한 형태의 이온 채널을 조절함에 의해 중추신경 세포의 흥분도를 조절하는 진세노사이드의 효과에 대하여도 보고된 바 있다. Recently, among the various effects of ginsenosides in vivo, the effects on the central nerve have been reported. The effects of ginseng on learning and memory have been found in animal models of learning and memory, especially the hippocampus known as the most important site for the formation of animal models damaged by scopolamine, weakening memory according to age, and memory formation. hippocampus) It is known that ginseng mitigates memory loss caused by nerve cell damage (Hsieh, MT et al. , Phytother.Res., 14: 375-377, 2000; Wesnes, KA, Ward et al. , Psychopharmacology (Berl), 152: 353-361, 2000). The effect of ginseng on this memory has been applied as a memory enhancer containing ginseng extract (Korean Patent Application No. 2000-4715). In addition, the effects of ginsenosides on the excitability of CNS by regulating various types of ion channels have been reported.
헌팅턴병은 신경퇴행성 유전질환으로 가장 흔히 관찰되는 유전성 신경계 질 환 형태들 중의 하나이다. 증상의 특징은 1) 성격변화, 우울증 등의 정신장애와 2) 운동 이상으로 나타난다. 발병률은 대체로 백만명당 4-5명으로 대체적인 발병 연령은 30-40대이지만 15세 이전, 심지어는 소아기에 시작되는 경우도 있다. 헌팅턴병은 뇌의 선조체(striatum) 및 기저핵(basal ganglia)과 대뇌피질의 신경세포의 선택적인 소실로 발병된다. 특히 미상핵(caudate nucleus)과 피각(putamen)의 두부에서 현저한 위축이 발견되며, 흔히 전두엽과 측두엽에서 중등도의 뇌회위축도 동반된다. 1993년 헌팅턴병의 원인유전자가 클로닝되었으며, 그 유전자 산물은 헌팅틴(huntingtin)이라는 단백질이다. 헌팅틴에 돌연변이가 일어났을 때 어떠한 기전으로 신경세포사가 일어나는가에 대한 메커니즘은 정확히 알려져있지 않으나 현재 활발한 연구가 진행중이다. 현재까지 알려진 바로는 캐스페이즈(caspase)라는 효소가 중요한 역할을 한다는점, 그리고 세포사를 일으킬 때 산화 스트레스가 중요한 역할을 할 것이라는 점이 있다(Gutekunst C.A., Norflus F., Curr. Opin. Neurol., 13:445-450, 2000). 현재 헌팅턴병에 대한 뚜렷한 치료법은 없는 실정이다. 환자의 운동 장애를 억제하기 위해 도파민 길항제인 할로페리돌(haloperidol)이 가장 널리 사용되고 있다. 기타 레세르핀(reserpine), 클로자핀(clozapine), 테트라바나진(tetrabanazine)과 같은 도파민 길항제는 헌팅턴병을 억제하는 효과가 있으나 졸음, 정좌 불능증, 만발성 운동이상증 등의 부작용이 크다. 할로페리돌을 비롯한 이러한 약들은 행동의 이상, 감정이 쉽게 변하는 등의 증상은 경감시킬 수 있으나 질환의 진행은 막지 못한다.Huntington's disease is one of the most common forms of hereditary nervous system disease seen as a neurodegenerative disorder. Symptoms are characterized by 1) mental disorders such as personality changes and depression, and 2) motor abnormalities. The incidence rate is usually 4-5 per million people, with an age of about 30 to 40 years, but often begins before age 15 and even in childhood. Huntington's disease is caused by selective loss of the brain's striatum and basal ganglia and cortical neurons. Significant atrophy is found in the head of the caudate nucleus and putamen, with moderate cerebral atrophy in the frontal and temporal lobes. In 1993, the causative agent of Huntington's disease was cloned and its gene product is a protein called huntingtin. The mechanism by which neuronal cell death occurs when mutations in huntingtin occur is not known, but active research is ongoing. To date, it is known that an enzyme called caspase plays an important role and that oxidative stress plays an important role in causing cell death (Gutekunst CA, Norflus F., Curr. Opin. Neurol., 13 : 445-450, 2000). There is currently no clear cure for Huntington's disease. The dopamine antagonist haloperidol (haloperidol) is most widely used to suppress the movement disorders of patients. Other dopamine antagonists, such as reserpine, clozapine, and tetrabanazine, have the effect of suppressing Huntington's disease, but have side effects such as drowsiness, dyspnea, and dyskinesia. These drugs, including haloperidol, can alleviate symptoms such as abnormal behavior and easily changing emotions, but do not prevent the progression of the disease.
한편, 3-니트로프로피온 산(3-nitropropionic acid; 3-NP)은 진균류에 오염 된 사탕수수 및 옥수수에서 발견된 화합물로, 동물 및 인간 모두에서 신경세포독성(neurotoxicity)을 나타내는 것으로 알려져 있다(Ming, Clin. Toxicol, 33:363-367, 1995; James L.F. et al., Am J. Vet. Res., 41:377-382, 1980). 만성적인 3-NP의 투여는 인간의 퇴행성 뇌질환의 일종인 헌팅톤병(Huntington's disease)과 유사한 임상적 증상을 유발시키는 것으로 알려져 있어, 3-NP는 헌팅톤병 연구를 위한 동물 모델에서 광범위하게 사용된다(Beal M.F. et al., J. Neurosci., 13:4181-4192, 1993). Meanwhile, 3-nitropropionic acid (3-NP) is a compound found in sugar cane and corn contaminated with fungi, and is known to exhibit neurotoxicity in both animals and humans (Ming). , Clin.Toxicol, 33: 363-367, 1995; James LF et al. , Am J. Vet.Res., 41: 377-382, 1980). Chronic 3-NP administration is known to cause clinical symptoms similar to Huntington's disease, a type of degenerative brain disease in humans, and 3-NP is widely used in animal models for Huntington's disease research. (Beal MF et al. , J. Neurosci., 13: 4181-4192, 1993).
그러나, 아직까지 만성적인 신경독성 손상에 의해 유도되는 점진적인 선조체의 변성(degeneration)에 대한 진세노사이드의 생체내 신경세포 보호효과에 대해서는 알려진 바 없다. However, there is no known in vivo neuroprotective effect of ginsenosides on the gradual degeneration of the striatum induced by chronic neurotoxic damage.
이에 본 발명자들은 3-NP에 의해 유도되는 선조체 신경독성, 운동신경 손상, 세포내 칼슘이온 농도 증가 및 미토콘드리아 손상을 홍삼 사포닌 혼합물이 유의적으로 억제하는 것을 확인함으로써 본 발명을 완성하였다.The present inventors completed the present invention by confirming that red ginseng saponin mixture significantly inhibited striatal neurotoxicity, motor neuron damage, increased intracellular calcium ion concentration and mitochondrial damage induced by 3-NP.
결국, 본 발명의 주된 목적은 홍삼 사포닌 혼합물을 함유하는 인지장애, 운동장애, 성격장애 또는 헌팅톤병의 예방 및 치료용 조성물을 제공하는데 있다.After all, the main object of the present invention is to provide a composition for the prevention and treatment of cognitive impairment, movement disorder, personality disorder or Huntington's disease containing red ginseng saponin mixture.
상기 목적을 달성하기 위하여, 본 발명은 진세노사이드 Rb1, Rb2, Rc, Rd, Re, Rf, Rg1, Rg2, Rg3, Ro 및 Ra를 필수적으로 함유하는 홍삼 사포닌 혼합물을 0.25~1.0 중량% 함유하는 3-NP(3-nitropropionic acid)에 의해 유발되는 선조체 신경 독성, 운동신경 손상, 세포내 칼슘이온 농도 증가 및 미토콘드리아의 손상에 의해 발생하는 인지장애, 운동장애, 성격장애 또는 헌팅톤병의 예방 또는 치료용 조성물을 제공한다.In order to achieve the above object, the present invention contains a ginsenoside Rb1, Rb2, Rc, Rd, Re, Rf, Rg1, Rg2, Rg3, Ro and Ra containing 0.25 to 1.0% by weight of a red ginseng saponin mixture essentially Prevention or treatment of cognitive, motor, personality, or Huntington's disease caused by striatal neurotoxicity caused by 3-NP (3-nitropropionic acid), motor neuron damage, increased intracellular calcium ion concentration, and mitochondrial damage It provides a composition for.
삭제delete
삭제delete
본 발명의 홍삼 진세노사이드는 통상적인 방법에 의하여 정제, 캅셀제, 과립제, 현탁제, 유제와 같은 단위투여형 또는 수회 투여형 제제로 제형화하여 사용될 수 있다. 그리고, 홍삼은 오랫동안 생약 및 식용으로 쓰여 왔던 약재로서 이들로부터 추출 및 분리된 본 발명의 화합물 역시 독성 및 부작용 등의 문제가 없다.The red ginseng ginsenosides of the present invention can be formulated into unit dosage forms or multiple dosage forms such as tablets, capsules, granules, suspensions, emulsions by conventional methods. In addition, red ginseng has been used for a long time as a herbal medicine and edible medicines of the present invention extracted and separated from them also have no problems such as toxicity and side effects.
삭제delete
본 발명의 홍삼 추출물 또는 진세노사이드 화합물들을 함유하는 조성물은 목적하는 바에 따라 비경구 투여하거나 경구 투여할 수 있으며, 본 발명에 의한 조성물의 바람직한 투여량은 환자의 상태 및 체중, 질병의 정도, 약물형태, 투여경로 및 기간에 따라 다르지만, 당업자에 의해 적절하게 선택될 수 있다. 그러나, 바람직한 효과를 위해서, 본 발명의 홍삼 추출물 또는 진세노사이드는 1일 12.5~250 mg/kg으로, 바람직하게는 12.5~200 mg/kg으로 투여하는 것이 좋다. 투여는 하루에 한번 투여할 수도 있고, 수회 나누어 투여할 수도 있다. 상기 투여량은 어떠한 면으로든 본 발명의 범위를 한정하는 것은 아니다.The composition containing the red ginseng extract or ginsenoside compounds of the present invention can be parenterally or orally administered as desired, and the preferred dosage of the composition according to the present invention is the condition and weight of the patient, the extent of the disease, the drug. Depending on the form, route of administration, and duration, it may be appropriately selected by those skilled in the art. However, for the desired effect, the red ginseng extract or ginsenoside of the present invention is preferably administered at 12.5 to 250 mg / kg, preferably at 12.5 to 200 mg / kg. Administration may be administered once a day or may be divided several times. The dosage does not limit the scope of the invention in any aspect.
본 발명은 또한, 상기 홍삼 추출물 또는 진세노사이드와 식품학적으로 허용 가능한 식품보조 첨가제를 포함하는, 인지장애, 운동장애, 성격장애 또는 헌팅톤병의 예방 효과를 나타내는 건강기능식품을 제공한다.The present invention also provides a health functional food comprising the red ginseng extract or ginsenoside and food supplements that can be used as a food supplement, which has a prophylactic effect of cognitive disorder, movement disorder, personality disorder or Huntington's disease.
본 발명에 있어서, 상기 홍삼 사포닌 혼합물은 인지장애, 운동장애, 성격장애 또는 헌팅톤병의 예방 효과를 목적으로 식품 또는 음료에 첨가될 수 있다. 이 때, 식품 또는 음료 중의 상기 홍삼 사포닌 혼합물의 양은 전체 식품 중량의 0.25~1.0중량%로 가할 수 있으며, 건강 음료 조성물은 100 ㎖를 기준으로 0.02~5 g, 바람직하게는 0.3~1 g의 비율로 가할 수 있다.In the present invention, the red ginseng saponin mixture may be added to food or beverage for the purpose of preventing cognitive impairment, movement disorder, personality disorder or Huntington's disease. At this time, the amount of the red ginseng saponin mixture in the food or beverage may be added at 0.25 to 1.0% by weight of the total food weight, the health beverage composition is 0.02 to 5 g, preferably 0.3 to 1 g based on 100 ml Can be added.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하고자 한다. 이들 실시예는 오로지 본 발명을 예시하기 위한 것으로서, 본 발명의 범위가 이들 실시예에 의해 제한되는 것으로 해석되지는 않는 것은 당업계에서 통상의 지식을 가진 자에게 있어서 자명할 것이다.Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention, it will be apparent to those skilled in the art that the scope of the present invention is not to be construed as being limited by these examples.
실시예 1: 실험재료 및 실험동물 Example 1 Experimental Materials and Experimental Animals
<1-1> 약물<1-1> drugs
하기 도 1에 본 발명의 대표적인 8개의 진세노사이드를 나타내었으며, 홍삼(Red ginseng) 전체 사포닌 혼합물(이하 'GTS'라 칭함)은 한국인삼공사(Korea Ginseng Corporation, Taejon, Korea)으로부터 분양 받았고, GTS 성분 중에는 Rb1(17.1%), Rb2(9.07%), Rc(9.65%), Rd(8.26%), Re(9%), Rf(3%), Rg1(6.4%), Rg2(4.2%), Rg3(3.8%), Ro(3.8%), Ra(2.91%) 및 다른 소수의 진세노사이드가 포함되어 있다. GTS는 사용하기 전에 bath medium 또는 식염수로 희석하여 사용하였으며, 3-NP(3-nitropropionic acid) 및 다른 시약들은 분석등급이며, 시그마사(St. Louis, MO)로부터 구입하여 사용하였다.1 shows eight representative ginsenosides of the present invention, and the red ginseng saponin mixture (hereinafter, referred to as 'GTS') was distributed from Korea Ginseng Corporation (Korea Ginseng Corporation, Taejon, Korea). Among the GTS components, Rb1 (17.1%), Rb2 (9.07%), Rc (9.65%), Rd (8.26%), Re (9%), Rf (3%), Rg1 (6.4%) and Rg2 (4.2%) , Rg3 (3.8%), Ro (3.8%), Ra (2.91%) and a few other ginsenosides. GTS was diluted with bath medium or saline prior to use, and 3-NP (3-nitropropionic acid) and other reagents were of analytical grade and purchased from Sigma (St. Louis, MO).
<1-2> 약물 투여 및 TTC 염색<1-2> Drug Administration and TTC Staining
수컷 스프라구-도울리계 랫트(300-350 g)를 사용하였으며, 온도, 습도 및 12시간 명암주기(AM. 7:00에 점등)가 조절되는 사육실에서 케이지 당 네 그룹으로 나누어 사육하였다. 45 마리의 랫트들을 임으로 식염수 투여군, 3-NP 단독 투여군 또는 GTS + 3-NP 투여군으로 나누었으며, 식이와 물은 자유로이 공급되도록 하였다. 대조군은 식염수만 투여하였으며, 식염수에 녹인 GTS는 27일 동안 12시간 간격으로 하루에 2번 50 ㎎/㎏의 투여량으로 랫트에 투여하였다. GTS는 3-NP를 투여(27일 동안 3일에 한번, 10 ㎎/㎏의 투여량)하기 전, 3일 동안 미리 투여하였으며, 3-NP 분말은 식염수에 녹여 사용하였고, pH는 수산화나트륨으로 7.4로 조정하였다. 3-NP를 3~4회 투여 후, 실험동물들은 급성적인 증상(ill)을 보였다. 투여횟수의 변화에 따라 실험동물들은 매츄(Mattews) 등의 연구에서 보인 바와 같은 증상을 나타내었다(Mattews et al., J. Neurosci., 18:156-163, 1998). 3-NP 투여 후, 실험동물을 3~4시간 동안 임상적으로 관찰하였으며, 급성적으로 증상을 보인 실험동물을 대조군 또는 홍삼 사포닌 혼합물-투여군에 상관없이 각 군으로부터 한 마리를 희생시켰다. 실험동물의 뇌를 신속히 제거한 다음 차가운 식염수 용액에 담가두었다가 랫트 뇌 매트릭스를 이용하여 2 ㎜ 간격으로 절개하였다. 절개된 절편은 37℃, 어두운 곳에서 5분동안 2% 2,3,5-트리페닐테트라졸리움 클로라이드(TTC) 용액으로 처리한 다음, TTC 용액을 제거하고 4% 파라포름알데히드(pH 7.4 in phosphate buffer)에 두었다. 컴퓨터에 기초한 이미지 분석(Image, NIH)을 이용하여, 병변(lesion)의 횡단면적을 합산하고 이 값에 절편 사이의 거리를 곱하여 병변 부피를 계산하였다. Male Sprague-Dawley rats (300-350 g) were used and housed in groups of four per cage in a control room controlled by temperature, humidity and a 12 hour contrast cycle (lit at AM 7:00). Forty-five rats were randomly divided into a saline-administered group, a 3-NP-only group, or a GTS + 3-NP-administered group. The diet and water were freely supplied. The control group was administered only saline, and GTS dissolved in saline was administered to rats at a dose of 50 mg / kg twice a day at 12 hour intervals for 27 days. GTS was pre-administered for 3 days before 3-NP administration (once every 3 days for 27 days, 10 mg / kg dose), 3-NP powder was dissolved in saline and the pH was adjusted to sodium hydroxide. Adjusted to 7.4. After three to four administrations of 3-NP, the experimental animals showed acute symptoms (ill). Depending on the number of doses, the animals exhibited the same symptoms as shown in the study of Matews et al. (Mattews et al. , J. Neurosci., 18: 156-163, 1998). After 3-NP administration, experimental animals were clinically observed for 3 to 4 hours, and acute symptoms of experimental animals were sacrificed from each group regardless of control or red ginseng saponin mixture-administered group. The brains of the experimental animals were quickly removed, soaked in cold saline solution, and cut in 2 mm intervals using a rat brain matrix. The incised sections were treated with 2% 2,3,5-triphenyltetrazolium chloride (TTC) solution at 37 ° C. in the dark for 5 minutes, then the TTC solution was removed and 4% paraformaldehyde (pH 7.4 in phosphate). buffer). Using computer-based image analysis (Image, NIH), the lesion volume was calculated by summing the cross-sectional areas of the lesions and multiplying this value by the distance between the sections.
실험결과, 홍삼 사포닌 혼합물은 3-NP로 유도된 선조체의 병변 부피를 용량-의존적으로 감소시킴을 확인할 수 있었으며, 홍삼 추출물(GE) 또한 3-NP 단독 투여군과 비교하여 선조체 병변 부피를 유의적으로 감소시킴을 확인할 수 있었다(도 2).As a result, the red ginseng saponin mixture was found to dose-dependently reduce the lesion volume of the 3-NP-induced striatum, and the red ginseng extract (GE) also significantly reduced the striatal lesion volume compared to the 3-NP alone group. It was confirmed that the reduction (Fig. 2).
<1-3> 선조체에 직접 투여한 말론산으로 유도된 선조체 병변에 대한 홍삼 사포닌 혼합물의 신경세포 보호효과 <1-3> Neuroprotective effect of red ginseng saponin mixture on malonic acid-induced striatal lesions administered directly to striatum
40 마리의 랫트를 임의로 식염수 투여군 및 GTS 투여군으로 나눈 다음, GTS는 12시간 간격으로 하루에 두 번, 50 ㎎/㎏의 투여량으로 말론산 투여 전, 3일 동안 미리 투여하였으며, GTS는 희생 전 일주일 동안 더 투여하였다. 0.01 M 농도의 말론산은 0.1 M PBS(pH 7.4)에 녹여 사용하였다. 실험동물은 마취 후 정위의 프 레임(stereotaxic frame)에 머리를 고정한 다음, 말론산을 다음의 정위와 대응되는 곳의 오른쪽 선조체에 주입하였다 : 브레그마 전방. 양귀간 선(interaural line) 2.3 ㎜ 아래의 노이즈 바 셋을 가진 A=0.8 ㎜; 중간선으로부터 측면, L=-2.8 ㎜; 및 정점의 아래 경막(dura), V=-6.0 mm. Forty rats were randomly divided into saline and GTS groups, and then GTS was administered twice a day at 12 hour intervals, at a dose of 50 mg / kg, prior to malonic acid for 3 days, and before GTS was sacrificed. It was administered for a week longer. Malonic acid at a concentration of 0.01 M was dissolved in 0.1 M PBS (pH 7.4). After anesthesia, the head was fixed to the stereotaxic frame after anesthesia, and then malonic acid was injected into the right striatum corresponding to the following position: Bregma anterior. A = 0.8 mm with a noise bar set below 2.3 mm of the interaural line; Lateral from midline, L = -2.8 mm; And dura below the vertex, V = -6.0 mm.
전체 1 ㎕를 주입 펌프가 설치된 10 ㎕의 하밀톤 실린지로 폴리에틸렌 튜브가 연결된 스테인레스 강철 캐뉼러를 통하여 0.25 ㎕/min의 속도로 주입하였다. 뇌를 제거한 후, 선조체의 병변 부피(striatal lesion volume)는 상기와 같이 TTC 염색으로 결정하였다.A total of 1 μl was injected at a rate of 0.25 μl / min through a stainless steel cannula with a polyethylene tube connected to a 10 μl Hamilton syringe equipped with an infusion pump. After removal of the brain, the striatal lesion volume of the striatum was determined by TTC staining as above.
실험결과, 도 3에 나타낸 바와 같이, 홍삼 사포닌 혼합물의 복강내 투여(100 ㎎/㎏)는 3-NP 단독 투여군과 비교하여 말론산으로 유도된 병변부피를 유의적으로 감소시킴을 확인할 수 있었다.As a result, as shown in Figure 3, it was confirmed that intraperitoneal administration of red ginseng saponin mixture (100 mg / kg) significantly reduced the volume of malonic acid-induced lesions compared to the 3-NP alone.
실시예 2: 홍삼 사포닌 혼합물의 신경세포 보호 효과 Example 2 of Red Ginseng Saponin Mixture Nerve cell protective effect
<2-1> 조직학적 관찰<2-1> Histological observation
홍삼 사포닌 혼합물의 신경세포보호 효과를 관상 절편의 크레실 바이올렛 염색을 통하여 확인하였다.The neuroprotective effect of the red ginseng saponin mixture was confirmed by Cresyl violet staining of coronal sections.
랫트들은 펜토발비탈(50 ㎖/㎏, i.p.)로 마취시킨 다음, 헤파린으로 응혈이 방지된 차가운 식염수(250 ㎖)를 이용하여 심장을 통하여 관류(perfusion)하였다. 식염수를 관류시킨 뒤 4% 파라포름알데히드(in 0.1M 인산염 완충액) 300 ㎖로 고정하였다. 뇌를 제거한 뒤 같은 고정액에서 하룻밤동안 전(前)고정한 다음, 30% 수크 로오스 용액으로 2일 동안 동결 방지하였다(cryoprotect). 뇌는 2-메틸부탄(이소펜탄)에서 동결하였으며, 조직화학적(histochemical) 및 면역조직화학적(immunohistochemical) 실험에 사용하기 위하여 -80℃에서 저장하였다. 브레그마 전방의 0-1.2 ㎜ 수준의 관상절편(40 ㎛ 두께)을 냉동미세절단기(cryostat, Leica, 1800, UK)로 잘라내었다. 크레실 바이올렛 염색을 하기 위하여 절편을 젤라틴-코팅된 슬라이드에 올려놓은 뒤 1% 크레실 바이올렛 아세테이트(시그마사, 미국) 용액을 이용하여 염색하였다. MADPH 다이포라아제 염색을 0.02M 인산염 완충액, 10 ㎎/㎖ NADPH(환원된 형태, 시그마) 및 1 ㎎/㎖ 니트로블루 테트라졸리움(시그마사, 미국)이 포함된 반응 혼합액으로 37℃, 어두운 곳에서 60분 동안 배양한 자유-부유하는 냉동미세절단된 절편에 수행하였다. Rats were anesthetized with pentovalbital (50 mL / kg, i.p.) and then perfused through the heart with cold saline (250 mL) protected from heparin coagulation. The saline was perfused and fixed with 300 ml of 4% paraformaldehyde (in 0.1 M phosphate buffer). The brains were removed and pre-fixed overnight in the same fixative and then cryoprotected for 2 days with 30% sucrose solution. Brains were frozen in 2-methylbutane (isopentane) and stored at −80 ° C. for use in histochemical and immunohistochemical experiments. Coronal sections (40 μm thick) at the 0-1.2 mm front of the bregma were cut out with a cryostat, Leica, 1800, UK. Sections were placed on gelatin-coated slides for cresyl violet staining and stained using 1% cresyl violet acetate (Sigma, USA) solution. MADPH diphorase staining was performed in a reaction mixture containing 0.02 M phosphate buffer, 10 mg / ml NADPH (reduced form, Sigma) and 1 mg / ml nitroblue tetrazolium (Sigma, USA) at 37 ° C. in the dark. Free-suspended frozen microcut sections were incubated for 60 minutes.
실험결과, 3-NP 단독 투여된 동물은 좌우대칭의 선조체 병변을 보인 반면, 홍삼 사포닌 혼합물과 3-NP를 함께 투여한 군에서는 선조체 양쪽 부분에서 약간의 병변만을 확인할 수 있었다(도 4). 또한, 홍삼 사포닌 혼합물은 3-NP 유도된 세포 손실을 유의적으로 감소시켰으며, 홍삼 사포닌 혼합물 + 3-NP 투여군에서는 긴 경로의 신경세포간 신경전달 형태를 갖는 NADPH 다이포라아제를 관찰할 수 있었다(도 4의 A 내지 F).As a result, animals treated with 3-NP alone showed symmetrical striatum lesions, while in the group administered with red ginseng saponin mixture and 3-NP, only slight lesions were found at both sides of the striatum (FIG. 4). In addition, the red ginseng saponin mixture significantly reduced 3-NP-induced cell loss, and the red ginseng saponin mixture + 3-NP group was able to observe NADPH diphorase with long-path neuronal neurotransmitter. (A to F of FIG. 4).
<2-2> 면역조직화학적 관찰<2-2> Immunohistochemical Observation
면역조직화학적 실험을 위하여, 관상절편을 마우스 GFAP(glial fibrially acidic protein- 이 단백질은 신경세포가 아닌 glia 세포에만 존재함) 단일세포 항체(시그마사, 1:1000 희석), 0.3% 트리톤 X-100, 0.5 ㎎/㎖ BSA(bovine serun albumin) 및 1.5% 정상 염소 혈청이 포함된 PBS로 4℃에서 하룻밤동안 배양한 다음, PBS 용액으로 각각 5분동안 세차례 세척하고 염소 항-마우스 Ig G(벡터, 1:200)로 1시간동안 배양한 후, 아비딘-바이오틴-퍼옥시다아제(avidin-biotin-peroxidase) 착물(벡터, 1:200)로 실온에서 1시간동안 배양하였다. 관상절편을 0.005% 3,3′-디아미노벤지딘-4HCl(DAB) 및 0.01% H2O2로 10분동안 발색시킨 다음, 헤마톡실린(hematoxylin)으로 염색하였다.For immunohistochemical experiments, the coronal sections were mouse GFAP (glial fibrially acidic protein), which is present only in glia cells, not neurons. Single-cell antibodies (Sigma, 1: 1000 dilution), 0.3% Triton X-100 , Incubated overnight at 4 ° C. in PBS containing 0.5 mg / ml BSA (bovine serun albumin) and 1.5% normal goat serum, and then washed three times with PBS solution for 5 minutes each and chlorine anti-mouse Ig G (vector, 1: 200) and then incubated with avidin-biotin-peroxidase complex (vector, 1: 200) for 1 hour at room temperature. Coronal sections were developed with 0.005% 3,3′-diaminobenzidine-4HCl (DAB) and 0.01% H 2 O 2 for 10 minutes and then stained with hematoxylin.
실험결과, 3-NP는 병변 코어(core)에서 GFAP 면역반응의 결여를 야기하였으며(도 4의 H), 홍삼 사포닌 혼합물은 대조군과 마찬가지로 3-NP에 의해 야기된 GFAP 양성 세포의 손실을 즉 성상글리아세포(astrocytes)와 같은 글리아(glia) 세포들을 감소시킴을 확인할 수 있었다(도 4의 G 및 I).As a result, 3-NP resulted in a lack of GFAP immune response in the lesion core (H of FIG. 4), and the red ginseng saponin mixture, as in the control group, showed loss of GFAP positive cells caused by 3-NP, i.e. Glia cells such as glia cells (astrocytes) was found to reduce (G and I of Figure 4).
실시예 3: 행동 테스트 및 생존 Example 3: Behavioral Testing and Survival
45 마리의 실험동물을 임으로 대조군, 3-NP 단독 또는 GTS 투여군, 및 3-NP 투여군으로 나누었으며, 약물 투여 순서는 실시예 1-2와 같은 방법으로 수행하였다. 각 군의 실험동물들은 식염수 또는 3-NP를 3일에 한번씩 투여(Keene et al., Exp. Neurol., 171:351-360, 2002)한 후, 지각 능력을 3-4시간동안 관찰하였다. 실험동물을 각 실험군으로 분리하기 전에, 50 ㎝ 디스크에 의해 네부분으로 분획된 6.0 ㎝ 직경(Ugo Basile, Italy)의 바(bar)로 구성된 장치에서 3일동안 최대 180초동안 연속적으로 매일 세 차례 훈련시켰으며, 성공하지 못한 동물들은 다음 실험에 서 배제하였다. 바는 가속도에 따라 5회전/분에서 25회전/분으로 회전하고, 각각의 시도에서 실험동물이 하강 전 장치에서 소비할 수 있는 시간을 시도 최대 레턴시(latency) 180초로 결정하여 측정하였다. 세 번의 결과를 평균으로 하여 기록하였다. 약물 투여후, 생존 실험에서 임의적으로 60마리의 실험동물을 대조군, 3-NP 단독 투여군 또는 GTS + 3NP 투여군으로 나누었으며, 약물 투여 순서는 상기 실시예 1-2와 같이 수행하였다. 각 군의 동물은 이른 아침 및 늦은 오후, 하루에 2회 관찰하였다. 실험동물의 행동 및 먹이를 섭취하는 능력을 면밀히 모니터링하였으며, 이를 동물을 안락사시키는 시기를 결정하는 자료로 채택하였다. 희생을 위한 기준은 실험동물이 사지 마비되어 등을 곧추 세우지 못하는 시점으로 하였다.45 experimental animals were randomly divided into a control group, 3-NP alone or GTS-administered group, and 3-NP-administered group, and the drug administration sequence was performed in the same manner as in Example 1-2. Animals in each group were treated with saline or 3-NP once every three days (Keene et al. , Exp. Neurol., 171: 351-360, 2002) and observed perceptual ability for 3-4 hours. Before separating the animals into each experimental group, three times daily for up to 180 seconds for three days in a device consisting of a 6.0 cm diameter (Ugo Basile, Italy) bar divided into four portions by a 50 cm disc. Trained and unsuccessful animals were excluded in the next experiment. The bar was rotated from 5 rotations / minute to 25 rotations / minute according to the acceleration, and the time that the experimental animal can spend on the device before descent in each trial was determined by determining the trial maximum latency of 180 seconds. Three results were recorded as averages. After drug administration, 60 experimental animals were randomly divided into a control group, 3-NP alone group, or GTS + 3NP group in the survival experiment, and the drug administration sequence was performed as in Example 1-2. Animals in each group were observed twice daily, early morning and late afternoon. The behavior of animals and their ability to eat food were closely monitored and were used as data to determine when animals were euthanized. The criterion for sacrifice was when the test animal was paralyzed and failed to straighten its back.
실험결과, 3-NP 단독 처리군은 세 차례의 3-NP 투여 후, 행동 장애(rota-rod로 측정)를 보이기 시작하였으며, 3-NP의 투여 횟수가 증가할 수록 행동적 기능장애가 악화되었다(도 5). 반면, 홍삼 사포닌 혼합물 처리군은 향상된 로타-로드 수행능력을 보였으며 3-NP 단독 처리군과 비교하여 향상된 지각능력을 나타내었다(도 5). 또한, 홍삼 사포닌 혼합물 처리군이 3-NP 단독 처리군에 비하여 높은 생존율을 나타내었다(도 6).As a result, the 3-NP alone treated group showed behavioral disorders (measured by rota-rod) after three administrations of 3-NP, and the behavioral dysfunction worsened as the number of administration of 3-NP increased. 5). On the other hand, the red ginseng saponin mixture treatment group showed improved rota-rod performance and improved perception ability compared to the 3-NP alone treatment group (FIG. 5). In addition, the red ginseng saponin mixture treatment group showed a higher survival rate than the 3-NP alone treatment group (Fig. 6).
실시예 4: 홍삼 사포닌 혼합물의 신경보호 효과의 메카니즘 규명 Example 4 Identification of Neuroprotective Effect of Red Ginseng Saponin Mixture
생체내(in vivo) 3-NP 투여에 의하여 유도된 뇌 선조체 독성에 대한 홍삼 사포닌 혼합물의 신경보호 효과의 메카니즘을 규명하기 위하여 하기와 같이 실험하였다. 3-NP의 처리는 배양된 뇌 선조체의 신경세포(neurons) 및 성상글리아세포(astrocytes)에서 서서히 세포내 Ca2+ 농도를 증가시키며, 최종적으로 세포사를 유발한다고 보고되었다(Fukuda et al., Neuroscience, 81:141-149, 1998). 본 발명의 홍삼 사포닌 혼합물이 배양된 선조체 세포들(neurons 및 astrocytes)에서 3-NP에 의해 유도된 Ca2+ 농도의 증가를 저해할 수 있는지를 실험하였다.In order to investigate the mechanism of neuroprotective effect of red ginseng saponin mixture on brain striatum toxicity induced by 3-NP administration in vivo , the following experiments were performed. Treatment of 3-NP has been reported to slowly increase intracellular Ca 2+ concentrations in neurons and astrocytes of cultured brain striatum and finally induce cell death (Fukuda et al. , Neuroscience , 81: 141-149, 1998). It was tested whether the red ginseng saponin mixture of the present invention could inhibit the increase of Ca 2+ concentration induced by 3-NP in cultured progenitor cells (neurons and astrocytes).
<4-1> 랫트 선조체 세포(striatal cell)의 배양 준비<4-1> Culture Preparation of Rat Striatal Cells
선조체 신경세포(strial neurons) 및 신경아교세포(glial cell)의 공배양은 크니(Keene) 등의 방법(Keene et al., Exp. Neurol., 171:351-360, 2001)을 변형하여 수행하였다. Coculture of striatal neurons and glial cells was performed by modifying the method of Keene et al. (Keene et al. , Exp. Neurol., 171: 351-360, 2001).
즉, 선조체를 16~18 일령의 암컷 스프라구-도울리계 랫트에서 분리한 다음, 0.25% 트립신(in HBSS) 용액으로 37℃에서 15분 동안 배양하였다. 세포는 fire-polished 파스퇴르 피펫으로 분쇄(trituration)하여 기계적으로 분리하였으며, 35㎜ 배양접시 또는 6-웰 플레이트의 폴리-L-리신-코팅된 커버슬립에서 배양하였다. 세포는 37℃의 95%의 습한 공기 및 5% 이산화탄소에서 10% FBS, 2% B-27 서플러먼트(supplement), 20 mM 글루코오스, 26.2 mM 소디움 바이카보네이트, 100 U/㎖ 페니실린 및 100 ㎍/㎖ 스트렙토마이신이 포함된 DMEM(Dullbecco's modified Eagle's medium; Life Technologies, Inc. Grand Island, NY, USA) 배지에서 유지하였다. 실험은 시험관내에서 7-15일 배양된 세포를 이용하여 수행하였다. 3-NP는 식염수에서 100 × stocks 으로 준비하였으며, NaOH로 pH 7.4가 되도록 중화하였다. 사용된 3-NP의 최종 농도는 5 mM 이었다.That is, the striatum was separated from female Sprague-Dawley rats of 16 to 18 days of age, and then incubated with 0.25% trypsin (in HBSS) solution at 37 ° C. for 15 minutes. Cells were mechanically isolated by trituration with a fire-polished Pasteur pipette and cultured in poly-L-lysine-coated coverslips in 35 mm petri dishes or 6-well plates. Cells were 10% FBS, 2% B-27 supplement, 20 mM glucose, 26.2 mM sodium bicarbonate, 100 U / ml penicillin and 100 μg / in 95% humid air and 5% carbon dioxide at 37 ° C. DMEM (Dullbecco's modified Eagle's medium; Life Technologies, Inc. Grand Island, NY, USA) medium containing ml streptomycin was maintained. Experiments were performed using cells cultured 7-15 days in vitro. 3-NP was prepared at 100 × stocks in saline and neutralized to pH 7.4 with NaOH. The final concentration of 3-NP used was 5 mM.
<4-2> 세포내 Ca<4-2> intracellular Ca 2+2+ 농도 측정 Concentration measurement
아세톡시메틸-에스테르 형태의 푸라-2(fura-2/AM; Molecular probes, Eugene, OR)를 형광성 Ca2+ 표지물로 사용하였으며, 150 mM NaCl, 5 mM KCl, 1 mM MgCl2, 2 mM CaCl2, 10 mM HEPES 및 10 mM 글루코오스로 구성된 HEPES 완충용액(pH 7.4)에서 5 μM 푸라-2/AM와 0.001% 플루로닉(Pluronic) F-127와 함께 신경세포를 실온에서 40 ~ 60분 동안 배양하여 표지한 후, HEPES 완충용액으로 세척한 다음 도립 현미경(inverted microscope, Olympus, Japan)을 사용하여 세포내 칼슘 이온 농도 변화를 측정하였다.Fura-2 (Fura-2 / AM; Molecular probes, Eugene, OR) in acetoxymethyl-ester form was used as fluorescent Ca 2+ label, 150 mM NaCl, 5 mM KCl, 1 mM MgCl 2 , 2 mM CaCl Neurons were cultured for 40 to 60 minutes at room temperature with 5 μM Pura-2 / AM and 0.001% Pluronic F-127 in HEPES buffer (pH 7.4) consisting of 2 , 10 mM HEPES and 10 mM glucose. After incubation and labeling, the cells were washed with HEPES buffer, and then intracellular calcium ion concentration was measured by using an inverted microscope (Ollympus, Japan).
구체적으로, 크세논 아크 램프를 비추어 컴퓨터 제어 필터 휠(Sutter Instruments, CA)에 의해 여기 파장(340 및 380nm)을 선택적으로 세포에 노출시켰다. 매 2-5초간격으로 데이터를 얻었으며, 빛이 노출되는 사이에 셔터를 설치하여 세포에 주는 광독성(phototoxicity)을 방지하였다. 515 nm의 고대역 통과 여과기(long-pass filter)를 통과하여 들어온 방출 형광(emitter fluorescence light)은 냉각 CCD 카메라를 지나 디지털 형광 분석기에 의해 비율값과 세포내 유리 Ca2+ 농도([Ca2+]i) 값으로 전환하였다. 모든 영상 데이터와 분석은 유니버셜 이미지 소프트웨어(West Chester, PA)를 이용하여 분석하였다(Kim et al., Biochem. Biophy. Res. Comm. 296, 247-254, 2002). Specifically, excitation wavelengths (340 and 380 nm) were selectively exposed to cells by a computer controlled filter wheel (Sutter Instruments, CA) in the light of a xenon arc lamp. Data were taken every 2-5 seconds, and shutters were placed between exposures of light to prevent phototoxicity to the cells. Emitter fluorescence light coming in through a 515 nm long-pass filter was passed through a cooled CCD camera and passed by a digital fluorescence analyzer to a ratio value and intracellular free Ca 2+ concentration ([Ca 2+]). i ) were converted to values. All image data and analyzes were analyzed using Universal Image Software (West Chester, PA) (Kim et al. , Biochem. Biophy. Res. Comm. 296, 247-254, 2002).
실험결과, 홍삼 사포닌 혼합물은 3-NP에 의해 유도된 Ca2+ 농도의 증가를 유의적으로 억제시킴을 확인할 수 있었다 ( *P < 0.001, 도 7).As a result, it was confirmed that the red ginseng saponin mixture significantly inhibited the increase of Ca 2+ concentration induced by 3-NP ( * P <0.001, FIG. 7).
<3-3> 미토콘드리아 막포텐셜의 측정<3-3> Measurement of mitochondrial membrane potential
3-NP에 의해 유도된 Ca2+ 농도의 증가는 미토콘드리아의 기능장애를 유발할 수 있으므로, 하기와 같이 홍삼 사포닌 혼합물의 존재 및 부존재 하에서 3-NP 유도된 미토콘드리아막의 포텐셜 변화를 측정하였다.Since the increase of Ca 2+ concentration induced by 3-NP may cause mitochondrial dysfunction, the potential change of 3-NP induced mitochondrial membrane in the presence and absence of red ginseng saponin mixture was measured as follows.
미토콘드리아막의 포텐셜(△Ψm)은 △Ψm-감각 염색법, 로드아민123(Rh123; Molecular probes, Eugene, OR)(Kannurpatti et al., Neurochem. Int., 44:361-369, 2004)을 이용하여 모니터링하였다. Mitochondrial membrane potential (△ Ψ m) is △ Ψ m - sense staining, loaded amine 123 (Rh123; Molecular probes, Eugene , OR): using a (Kannurpatti et al, Neurochem Int, 44 361-369, 2004...) Monitoring.
즉, 세포를 실온에서 5 ㎍/㎖의 염색제로 15분 동안 어두운 곳에서 배양하고 로딩(loading)한 다음, HEPES 완충용액으로 세 차례 세척하였다. 홍삼 사포닌 혼합물 + 3-NP 투여군은 홍삼 사포닌 혼합물(100 ㎍/㎖)을 홍삼 사포닌 혼합물과 3-NP를 함께 처리하기 전 1분 동안 미리 처리하였다. 크세논 아크 램프를 비추어 컴퓨터 제어 필터 휠(Sutter Instruments, CA)에 의해 여기 파장(490 nm)을 선택적으로 세포에 노출시켰다. 방출 형광(emitter fluorescence light)은 537 nm의 고대역 통과 여과기(long-pass filter)를 통과하여 플레임 변형 냉각 CCD(frame transfer cooled CCD) 카메라에 반사되었다. 매 2-5초 간격으로 데이터를 얻었으며, 빛이 노출되는 사이에 셔터를 설치하여 세포에 주는 광독성(phototoxicity)을 방지하였다. 모든 영상 데이터와 분석은 유니버셜 이미지 소프트웨어(West Chester, PA)를 이용하여 분석하였다. 시작점의 형광강도(fluorescent intensity)는 1로 나타내었다.That is, the cells were incubated for 15 minutes in the dark with 5 μg / ml of staining agent at room temperature, loaded, and then washed three times with HEPES buffer. Red ginseng saponin mixture + 3-NP administration group was previously treated with red ginseng saponin mixture (100 ㎍ / ㎖) for 1 minute before the red ginseng saponin mixture and 3-NP. The xenon arc lamp was lighted to selectively expose the excitation wavelength (490 nm) to the cells by a computer controlled filter wheel (Sutter Instruments, Calif.). Emitter fluorescence light was passed through a 537 nm long-pass filter and reflected on a frame transfer cooled CCD camera. Data was taken every 2-5 seconds, and shutters were placed between exposures of light to prevent phototoxicity to cells. All image data and analyzes were analyzed using Universal Image Software (West Chester, PA). Fluorescent intensity of the starting point is represented by 1.
실험결과, 3-NP로 인한 미토콘드리아막의 포텐셜의 감소(0.86 ± 0.01)를 홍삼 사포닌 혼합물이 존재할 경우에는 0.92 ± 0.01로 유의적으로 증가시킴을 확인할 수 있었다( *P < 0.001, 도 8).As a result, it was confirmed that the potential of the mitochondrial membrane due to 3-NP (0.86 ± 0.01) was significantly increased to 0.92 ± 0.01 in the presence of red ginseng saponin mixture ( * P <0.001, Figure 8).
실시예 4: 3-NP 유도된 선조체 세포독성에 대한 홍삼 사포닌 혼합물의 저해효과 Example 4 Inhibitory Effect of Red Ginseng Saponin Mixture on 3-NP Induced Progenitor Cytotoxicity
<4-1> MTT 테스트법<4-1> MTT test method
신진대사 저해 또는 미토콘드리아 기능장애로 인한 세포사는 MTT (3-(4,5-디메틸티아졸-2-일)-2,5-디페닐테트라졸리움 브로마이드)법을 이용하여 결정하였다(Mosmann, J. Immunol Methods, 16:55-63, 1983). Cell death due to metabolic inhibition or mitochondrial dysfunction was determined using MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide) method (Mosmann, J. Immunol Methods, 16: 55-63, 1983).
구체적으로, 세포를 혈청-유리 배지(serum-free medium)로 세척한 다음, 5 mM 3-NP 단독 또는 다양한 농도의 GTS의 존재하에 37℃에서 24시간동안 배양하였다. 세포를 3-NP로 처리하기 전에 GTS로 5분 동안 전처리한 다음 혈청-유리 배양 배지에서 24시간동안 배양한 후, 배양액은 세포 생존(cell viability) 기간(extent)동안 공급하였다. 세포 생존율은 MTT 테스트(test) 법을 이용하여 살아있는 세포에 존재하는 디히드로게나제 활성을 검출하여 측정하였다. PBS에서 MTT 용 액(1 ㎎/㎖)의 표준용액(aliquot) 50 ㎕를 세포에 직접적으로 첨가한 다음, 포르마잔에 신진대사시키기 위하여 MTT 용액으로 4시간 동안 배양하였다. 배양 후, 상청액은 흡출시키고 포르마잔을 용해시키기 위하여 100 ㎕ DMSO를 가하였다. OD(optical density)를 560 nm 파장에서 automated spectrophotometric plate reader로 측정하였으며, 대조군과 비교한 상대적인 세포 생존율을 결정하였다.Specifically, the cells were washed with serum-free medium and then incubated for 24 hours at 37 ° C. in the presence of 5 mM 3-NP alone or in various concentrations of GTS. Cells were pretreated for 5 minutes with GTS prior to treatment with 3-NP and then incubated for 24 hours in serum-free culture medium, and the cultures were fed for a period of cell viability. Cell viability was determined by detecting the dehydrogenase activity present in living cells using the MTT test method. 50 μl of MTT solution (1 mg / ml) in PBS was added directly to the cells, and then incubated with MTT solution for 4 hours to metabolize in formazan. After incubation, the supernatant was aspirated and 100 μl DMSO was added to dissolve the formazan. Optical density (OD) was measured with an automated spectrophotometric plate reader at a wavelength of 560 nm, and the relative cell viability compared to the control group was determined.
실험결과, 도 9에 나타낸 바와 같이 3-NP 처리에 의해 세포 생존율이 51 ± 2%까지 감소하였으나, 홍삼 사포닌 혼합물을 전(前)처리한 경우 용량-의존적으로 세포 생존율이 유의적으로 증가됨을 관찰할 수 있었다. 세포 생존율은 홍삼 사포닌 혼합물 3, 10, 30 및 100㎍/㎖에 대하여 각각 68.7 ± 1.6, 77.8 ± 1.9, 84.4 ± 2.0 및 92.1 ± 2.1%로 증가하였으며, 홍삼 사포닌 혼합물 1 ㎍/㎖에서는 유의적인 효과가 관찰되지 않았다.As a result, as shown in Figure 9, the cell survival rate was reduced to 51 ± 2% by 3-NP treatment, but when the pre-treatment of red ginseng saponin mixture was pre-treated, the cell survival rate was significantly increased. Could. Cell viability was increased to 68.7 ± 1.6, 77.8 ± 1.9, 84.4 ± 2.0 and 92.1 ± 2.1% for red
<4-2> DNA 래더링(laddering) 측정<4-2> DNA Laddering Measurement
DNA 래더(ladder)를 측정하기 위하여, PBS(pH 7.4)에서 배양된 세포를 수확한 다음 4℃, 12000 × g에서 5분 동안 원심 분리하였다. 키트(GeneAllTM)를 이용하여 대조군, 3-NP 단독 또는 GTS군 및 3-NP 투여군에서 배양된 세포로부터 DNA를 추출하였다. DNA 단편은 50V에서 1시간동안 1.4% 아가로스 겔 전기이동법으로 측정하였으며, 에티디움 브로마이드(ethidium bromide)로 염색시켰다. To measure the DNA ladder, cells cultured in PBS (pH 7.4) were harvested and centrifuged for 5 minutes at 12000 × g at 4 ° C. DNA was extracted from cells cultured in a control group, 3-NP alone or GTS group and 3-NP administration group using a kit (GeneAll ™ ). DNA fragments were measured by 1.4% agarose gel electrophoresis at 50V for 1 hour and stained with ethidium bromide.
실험결과, 홍삼 사포닌 혼합물은 3-NP에 의해 유도된 핵의 DNA 절편화를 저해시킴을 확인할 수 있었다(도 10).Experimental results, it was confirmed that the red ginseng saponin mixture inhibits DNA fragmentation of the nucleus induced by 3-NP (Fig. 10).
실시예 5: 데이터 분석 Example 5: Data Analysis
모든 데이터는 평균 ± S.E.M.으로 나타내었으며, 통계분석을 위하여 다중분석에 사용되는 Tukey' test 및 스튜던트 T 테스트에 의한 ANOVA 분석법을 사용하였다. All data are expressed as mean ± S.E.M., and ANOVA analysis by Tukey 'test and Student T test used for multiple analysis for statistical analysis.
이상으로 본 발명 내용의 특정한 부분을 상세히 기술하였는바, 당업계의 통상의 지식을 가진 자에게 있어서, 이러한 구체적 기술은 단지 바람직한 실시양태일 뿐이며, 이에 의해 본 발명의 범위가 제한되는 것이 아닌 점은 명백할 것이다. 따라서 본 발명의 실질적인 범위는 첨부된 청구항들과 그것들의 등가물에 의하여 정의된다고 할 것이다.The specific parts of the present invention have been described in detail above, and it is apparent to those skilled in the art that such specific descriptions are merely preferred embodiments, and thus the scope of the present invention is not limited thereto. something to do. Thus, the substantial scope of the present invention will be defined by the appended claims and their equivalents.
이상에서 상세히 설명한 바와 같이, 본 발명의 홍삼 사포닌 혼합물은 헌팅톤병(Huntington's disease)과 유사한 임상적 증상을 유발시키는 것으로 알려진 3-NP에 의해 유도되는 선조체 신경독성, 운동신경 손상, 세포내 칼슘이온 농도 증가 및 미토콘드리아 손상을 유의적으로 억제시키므로, 인지장애, 운동장애, 성격장애 또는 헌팅톤병의 예방 및 치료를 위한 의약품 또는 건강기능식품을 제공하는 효과가 있다.As described in detail above, the red ginseng saponin mixture of the present invention is the striatum neurotoxicity induced by 3-NP known to cause clinical symptoms similar to Huntington's disease, motor neuron damage, intracellular calcium ion concentration Since it significantly inhibits the increase and mitochondrial damage, there is an effect of providing a medicament or dietary supplement for the prevention and treatment of cognitive impairment, movement disorder, personality disorder or Huntington's disease.
Claims (13)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020040042590A KR100656888B1 (en) | 2004-06-10 | 2004-06-10 | Composition Comprising Ginsenosides Having Neuroprotective Effects against Brain Striatum Neurotoxicity |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020040042590A KR100656888B1 (en) | 2004-06-10 | 2004-06-10 | Composition Comprising Ginsenosides Having Neuroprotective Effects against Brain Striatum Neurotoxicity |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| KR20050117313A KR20050117313A (en) | 2005-12-14 |
| KR100656888B1 true KR100656888B1 (en) | 2006-12-13 |
Family
ID=37290737
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| KR1020040042590A KR100656888B1 (en) | 2004-06-10 | 2004-06-10 | Composition Comprising Ginsenosides Having Neuroprotective Effects against Brain Striatum Neurotoxicity |
Country Status (1)
| Country | Link |
|---|---|
| KR (1) | KR100656888B1 (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20170005216A (en) | 2015-07-01 | 2017-01-12 | 재단법인 지능형 바이오 시스템 설계 및 합성 연구단 | Method for screening activator of mitochondria activity |
| US20210100823A1 (en) * | 2018-04-23 | 2021-04-08 | Sheau-Long Lee | Use of ginsenoside m1 for treating huntington's disease |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2001139483A (en) | 1999-08-30 | 2001-05-22 | Japan Science & Technology Corp | Protecting agent for brain cell or nerve cell, consisting of ginseng |
| KR20020084311A (en) * | 2001-04-25 | 2002-11-07 | (주) 디지탈바이오텍 | Composition for preventing or treating involutional brain diseases comprising a hydrolysate of ginsenosides |
| US6579853B2 (en) | 1998-12-22 | 2003-06-17 | Japan Science And Technology Corporation | Brain cell or nerve cell-protective agents comprising ginsenoside Rb1 |
| KR20030059984A (en) * | 2002-01-05 | 2003-07-12 | 박명환 | A new process for enhancing the efficacies of Ginseng and the uses of the Ginseng extract |
| KR20030088417A (en) * | 2003-11-06 | 2003-11-19 | 주식회사 케이티앤지 | The memory increasing novel compound ginseno-jk from ginseng hairy roots and the manufacturing method of the ginseno-jk |
| KR20040036451A (en) * | 2002-10-26 | 2004-04-30 | 한국과학기술연구원 | Composition for inhibiting glutamate-mediated neurotoxicity comprising ginsenoside rg3 or ginsenoside rh2 |
-
2004
- 2004-06-10 KR KR1020040042590A patent/KR100656888B1/en not_active IP Right Cessation
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6579853B2 (en) | 1998-12-22 | 2003-06-17 | Japan Science And Technology Corporation | Brain cell or nerve cell-protective agents comprising ginsenoside Rb1 |
| JP2001139483A (en) | 1999-08-30 | 2001-05-22 | Japan Science & Technology Corp | Protecting agent for brain cell or nerve cell, consisting of ginseng |
| KR20020084311A (en) * | 2001-04-25 | 2002-11-07 | (주) 디지탈바이오텍 | Composition for preventing or treating involutional brain diseases comprising a hydrolysate of ginsenosides |
| KR20030059984A (en) * | 2002-01-05 | 2003-07-12 | 박명환 | A new process for enhancing the efficacies of Ginseng and the uses of the Ginseng extract |
| KR20040036451A (en) * | 2002-10-26 | 2004-04-30 | 한국과학기술연구원 | Composition for inhibiting glutamate-mediated neurotoxicity comprising ginsenoside rg3 or ginsenoside rh2 |
| KR20030088417A (en) * | 2003-11-06 | 2003-11-19 | 주식회사 케이티앤지 | The memory increasing novel compound ginseno-jk from ginseng hairy roots and the manufacturing method of the ginseno-jk |
Non-Patent Citations (2)
| Title |
|---|
| 1020040042590 - 710797 |
| 1020040042590 - 710798 |
Also Published As
| Publication number | Publication date |
|---|---|
| KR20050117313A (en) | 2005-12-14 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US9011937B2 (en) | Method of treating neurological conditions with extract of Nerium species or Thevetia species | |
| Chiu et al. | Formulated Chinese medicine Shaoyao Gancao Tang reduces NLRP1 and NLRP3 in Alzheimer’s disease cell and mouse models for neuroprotection and cognitive improvement | |
| JP6453791B2 (en) | Methods of treating neurological conditions with extracts of Oleander species or Thevetia species | |
| KR20140098811A (en) | Composition comprising vicenin-2 having a beneficial effect on neurological and/or cognitive function | |
| Bak et al. | Neuroprotective effects of 20 (S)-protopanaxadiol against glutamate-induced mitochondrial dysfunction in PC12 cells | |
| US20050170028A1 (en) | Polysaccharide extract from Lycium barbarum as neuroprotective agent against beta-amyloid peptide neurotoxicity | |
| Diling et al. | Docking studies and biological evaluation of a potential β-secretase inhibitor of 3-hydroxyhericenone F from Hericium erinaceus | |
| Lian et al. | Anticonvulsant and neuroprotective effects of ginsenosides in rats | |
| SG184742A1 (en) | Phase ii detoxification and antioxidant activity | |
| Miller et al. | A rat model of nerve agent exposure applicable to the pediatric population: the anticonvulsant efficacies of atropine and GluK1 antagonists | |
| Paidi et al. | Medha Plus–A novel polyherbal formulation ameliorates cognitive behaviors and disease pathology in models of Alzheimer’s disease | |
| Zhao et al. | Targeted metabolomics reveals the aberrant energy status in diabetic peripheral neuropathy and the neuroprotective mechanism of traditional Chinese medicine JinMaiTong | |
| Shan et al. | Classic famous prescription kai-xin-san ameliorates alzheimer’s disease via the wnt/β-catenin signaling pathway | |
| KR100656888B1 (en) | Composition Comprising Ginsenosides Having Neuroprotective Effects against Brain Striatum Neurotoxicity | |
| Wang et al. | Gleditsiae sinensis fructus Pills combined with Jujubae fructus attenuate chronic bronchitis via regulation of AGE-RAGE signaling pathway | |
| Wang et al. | Activation of PINK1-Parkin-dependent mitophagy in Tri-ortho-cresyl phosphate-treated Neuro2a cells | |
| EP4161550A1 (en) | Shan-zha for the treatment of depression and anxiety disorders | |
| Zhang et al. | Astragaloside IV protects against autoimmune myasthenia gravis in rats via regulation of mitophagy and apoptosis | |
| Jain | A Mechanistic Approach to Determination of Anti-diabetic activity of Calystegia sepium R. Br. Flowering plants in normal and Streptozotocin induced rats | |
| Carvalho et al. | Efficacy and safety of cumaru syrup as complementary therapy in mild persistent asthma: a double-blind, randomized, placebo-controlled study | |
| Huang et al. | Quantitative Proteomics Reveal the Protective Effects of the Combined Chinese Herbal Extracts Against Alzheimer's Disease via Modulating the Expression of Synapse-associated Proteins in Mouse Model | |
| WO2021085496A1 (en) | Composition, antioxidant agent, antisaccharification agent, neurite outgrowth promoter, and cognitive function improver | |
| Kakkar et al. | Alzheimer’s disease: potential of nanotailored nutraceuticals | |
| Srivastava et al. | Investigating Bacopa monnieri L. Therapeutic Potential for the Treatment of Neurological Diseases | |
| Li | Investigation on the Effect of Hyperglycemia-induced Methylglyoxal in Neurodegenerative Disorder |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A201 | Request for examination | ||
| E902 | Notification of reason for refusal | ||
| E902 | Notification of reason for refusal | ||
| E701 | Decision to grant or registration of patent right | ||
| GRNT | Written decision to grant | ||
| FPAY | Annual fee payment |
Payment date: 20121011 Year of fee payment: 7 |
|
| FPAY | Annual fee payment |
Payment date: 20140113 Year of fee payment: 8 |
|
| FPAY | Annual fee payment |
Payment date: 20141105 Year of fee payment: 9 |
|
| FPAY | Annual fee payment |
Payment date: 20160202 Year of fee payment: 10 |
|
| LAPS | Lapse due to unpaid annual fee |